Among patients who underwent primary surgery for non-small cell lung cancer (NSCLC), recurrent disease is frequent and cannot be accurately predicted solely from TNM stage and histopathological ...features. The aim of this study was to examine the association of tumor markers in pre-operative serum with recurrent disease.
Blood samples were collected prior to lung cancer surgery from 107 patients with stage I-III lung adenocarcinoma surgically treated at Lund University hospital, Lund, Sweden, between 2005 and 2011. The serum tumor markers Carcinoembryonic antigen (CEA), Neuron-specific enolase (NSE), Cancer antigen 125 (CA 125), Human epididymis protein 4 (HE4) and Carbohydrate antigen (CA 19-9) were analyzed retrospectively and clinical follow-up data were collected from patient charts. Forty (37%) patients were diagnosed with recurrent disease.
Sixty-eight (64%) patients had at least one elevated tumor marker prior to surgery. In analysis of disease-free survival (DFS), CA 125 and/or CA 19-9 were significantly associated with recurrent disease adjusted to stage and adjuvant treatment (hazard ratio 2.8, 95% confidence interval 1.4-5.7, p = 0.006).
High pre-operative serum CA 19-9 and/or CA 125 might indicate an increased incidence of recurrent disease in resectable lung adenocarcinomas.
Accurate histological classification and identification of fusion genes represent two cornerstones of clinical diagnostics in non-small cell lung cancer (NSCLC). Here, we present a NanoString gene ...expression platform and a novel platform-independent, single sample predictor (SSP) of NSCLC histology for combined, simultaneous, histological classification and fusion gene detection in minimal formalin fixed paraffin embedded (FFPE) tissue. The SSP was developed in 68 NSCLC tumors of adenocarcinoma (AC), squamous cell carcinoma (SqCC) and large-cell neuroendocrine carcinoma (LCNEC) histology, based on NanoString expression of 11 (CHGA, SYP, CD56, SFTPG, NAPSA, TTF-1, TP73L, KRT6A, KRT5, KRT40, KRT16) relevant genes for IHC-based NSCLC histology classification. The SSP was combined with a gene fusion detection module (analyzing ALK, RET, ROS1, MET, NRG1, and NTRK1) into a multicomponent NanoString assay. The histological SSP was validated in six cohorts varying in size (n = 11-199), tissue origin (early or advanced disease), histological composition (including undifferentiated cancer), and gene expression platform. Fusion gene detection revealed five EML4-ALK fusions, four KIF5B-RET fusions, two CD74-NRG1 fusion and three MET exon 14 skipping events among 131 tested cases. The histological SSP was successfully trained and tested in the development cohort (mean AUC = 0.96 in iterated test sets). The SSP proved successful in predicting histology of NSCLC tumors of well-defined subgroups and difficult undifferentiated morphology irrespective of gene expression data platform. Discrepancies between gene expression prediction and histologic diagnosis included cases with mixed histologies, true large cell carcinomas, or poorly differentiated adenocarcinomas with mucin expression. In summary, we present a proof-of-concept multicomponent assay for parallel histological classification and multiplexed fusion gene detection in archival tissue, including a novel platform-independent histological SSP classifier. The assay and SSP could serve as a promising complement in the routine evaluation of diagnostic lung cancer biopsies.
Neuroendocrine (NE) differentiation in prostate cancer has been correlated with a poor prognosis and hormone refractory disease. In a previous report, we demonstrated the presence of immunoreactive ...cytoplasmic hypoxia inducible factor 1 alpha (HIF1 alpha ), in both benign and malignant NE prostate cells. HIF1 alpha and HIF1 beta are two subunits of HIF1, a transcription factor important for angiogenesis. The aim of this study was to elucidate whether the cytoplasmic stabilization of HIF1 alpha in androgen independent NE differentiated prostate cancer is due to the presence of certain HIF1 alpha isoforms. We studied the HIF1 alpha isoforms present in 8 cases of benign prostate hyperplasia (BPH) and 43 cases of prostate cancer with and without NE differentiation using RT-PCR, sequencing analysis, immunohistochemistry and in situ hybridization. We identified multiple isoforms in both benign and malignant prostate tissues. One of these isoforms, HIF1 alpha 1.2, which was previously reported to be testis specific, was found in 86% of NE-differentiated prostate tumors, 92% of HIF1 alpha immunoreactive prostate tumors and 100% of cases of benign prostate hyperplasia. Immunohistochemistry and in situ hybridization results showed that this isoform corresponds to the cytoplasmic HIF1 alpha present in androgen-independent NE cells of benign and malignant prostate tissue and co-localizes with immunoreactive cytoplasmic HIF1 beta . Our results indicate that the cytoplasmic stabilization of HIF1 alpha in NE-differentiated cells in benign and malignant prostate tissue is due to presence of an HIF1 alpha isoform, HIF1 alpha 1.2. Co-localization of this isoform with HIF1 beta indicates that the HIF1 alpha 1.2 isoform might sequester HIF1 beta in the cytoplasm.
Disease recurrence in surgically treated lung adenocarcinoma (AC) remains high. New approaches for risk stratification beyond tumor stage are needed. Gene expression‐based AC subtypes such as the ...Cancer Genome Atlas Network (TCGA) terminal‐respiratory unit (TRU), proximal‐inflammatory (PI) and proximal‐proliferative (PP) subtypes have been associated with prognosis, but show methodological limitations for robust clinical use. We aimed to derive a platform independent single sample predictor (SSP) for molecular subtype assignment and risk stratification that could function in a clinical setting. Two‐class (TRU/nonTRU=SSP2) and three‐class (TRU/PP/PI=SSP3) SSPs using the AIMS algorithm were trained in 1655 ACs (n = 9659 genes) from public repositories vs TCGA centroid subtypes. Validation and survival analysis were performed in 977 patients using overall survival (OS) and distant metastasis‐free survival (DMFS) as endpoints. In the validation cohort, SSP2 and SSP3 showed accuracies of 0.85 and 0.81, respectively. SSPs captured relevant biology previously associated with the TCGA subtypes and were associated with prognosis. In survival analysis, OS and DMFS for cases discordantly classified between TCGA and SSP2 favored the SSP2 classification. In resected Stage I patients, SSP2 identified TRU‐cases with better OS (hazard ratio HR = 0.30; 95% confidence interval CI = 0.18‐0.49) and DMFS (TRU HR = 0.52; 95% CI = 0.33‐0.83) independent of age, Stage IA/IB and gender. SSP2 was transformed into a NanoString nCounter assay and tested in 44 Stage I patients using RNA from formalin‐fixed tissue, providing prognostic stratification (relapse‐free interval, HR = 3.2; 95% CI = 1.2‐8.8). In conclusion, gene expression‐based SSPs can provide molecular subtype and independent prognostic information in early‐stage lung ACs. SSPs may overcome critical limitations in the applicability of gene signatures in lung cancer.
What's new?
New tools are needed in order to improve risk stratification and therapy selection in early‐stage lung adenocarcinoma. Inherent differences in gene expression between adenocarcinoma subtypes could facilitate the development of such tools. The authors of this study derived platform‐independent, single‐sample predictors (SSP) of adenocarcinoma subtypes, based on gene expression. Derived SSPs successfully provided prognostic information in surgically treated stage I lung adenocarcinoma patients. The single‐sample classifier was readily translated into assays applicable to archival tissue, indicating clinical utility. The findings highlight the clinical relevance of transcriptional signatures and gene expression predictors in lung adenocarcinoma, warranting their further investigation and development.
This paper presents results from an evaluation of three previously presented methods for segmentation of cell nuclei in lung cytology samples scanned by whole-slide scanners. Whole-slide images from ...seven cases of end bronchial ultrasound-guided Tran bronchial needle aspiration samples were used for extracting a number of regions of interest, in which approximately 2700 cell nuclei were manually segmented to form the ground truth. The segmented cells included benign bronchial epithelium, lymphocytes, granulocytes, histiocytes and malignant epithelial cells. The best results were obtained with a method based upon adaptive thresholding and an added step of clustering for distinguishing between cytoplasm and cell nuclei. This method achieved a mean DICE-score of 0.81 and a sensitivity and specificity of 0.88 and 0.81 respectively. In addition, this method was by far the fastest method, with a mean processing time of 7.8 s per image (2 048 × 2 048 pixels per image). By further improvements, such as lowering the false positive rate and using parallel computing hardware, this method has the potential to form the first building block in a system for computerized screening of whole-slide images in lung cytology.
Neuroendocrine (NE) differentiation in prostate cancer has been correlated with a poor prognosis and hormone refractory disease. In a previous report, we demonstrated the presence of immunoreactive ...cytoplasmic hypoxia inducible factor 1alpha (HIF1alpha), in both benign and malignant NE prostate cells. HIF1alpha and HIF1beta are two subunits of HIF1, a transcription factor important for angiogenesis. The aim of this study was to elucidate whether the cytoplasmic stabilization of HIF1alpha in androgen independent NE differentiated prostate cancer is due to the presence of certain HIF1alpha isoforms. We studied the HIF1alpha isoforms present in 8 cases of benign prostate hyperplasia (BPH) and 43 cases of prostate cancer with and without NE differentiation using RT-PCR, sequencing analysis, immunohistochemistry and in situ hybridization. We identified multiple isoforms in both benign and malignant prostate tissues. One of these isoforms, HIF1alpha1.2, which was previously reported to be testis specific, was found in 86% of NE-differentiated prostate tumors, 92% of HIF1alpha immunoreactive prostate tumors and 100% of cases of benign prostate hyperplasia. Immunohistochemistry and in situ hybridization results showed that this isoform corresponds to the cytoplasmic HIF1alpha present in androgen-independent NE cells of benign and malignant prostate tissue and co-localizes with immunoreactive cytoplasmic HIF1beta. Our results indicate that the cytoplasmic stabilization of HIF1alpha in NE-differentiated cells in benign and malignant prostate tissue is due to presence of an HIF1alpha isoform, HIF1alpha1.2. Co-localization of this isoform with HIF1beta indicates that the HIF1alpha1.2 isoform might sequester HIF1beta in the cytoplasm.
Small cell lung carcinoma (SCLC) is extremely aggressive and frequently metastasizes widely in its early stage. Because tumor hypoxia is related to aggressive tumor behavior and the hypoxic ...adaptation of SCLC is poorly documented, we stained SCLC tumors arranged in a tissue microarray for hypoxia-inducible factor (HIF)-1α and HIF-2α proteins. We found an overall lack of HIF-2α protein expression, which was confirmed in large tumor sections. HIF-1α protein was strongly expressed in most tumors, frequently adjacent to necrotic regions. In concordance, cultured SCLC but not non–small cell lung carcinoma cells showed no or extremely low levels of HIF-2α mRNA and no HIF-2α protein at hypoxia. HIF-1α was stabilized after 4 hours at hypoxia, and its accumulation increased up to 96 hours. SCLC cells survived well and showed net proliferation and low cell death in modest (1% oxygen) and severe (0.1% oxygen) hypoxia. HIF-1α repression virtually did not influence cell death or viability despite reduced levels of hypoxia-inducible genes, such as BNIP3 and BNIP3L . At 1% oxygen no increased autophagy (LC3B-II activation) or NF-κB signaling were detected, whereas the unfolded protein response was activated at severe hypoxia. Our data indicate that HIFs are not exclusively required for SCLC cell survival at modest or severe hypoxia and that additional, yet uncharacterized, hypoxia-driven adaptation pathways may become activated.
Summary
Background Eosinophils are seen at sites of inflammation in diseases such as helminthic infestation, asthma, ulcerative colitis and some neoplastic diseases. They are also associated with ...connective tissue remodelling, for example in longstanding asthma. In the present study, we investigated whether eosinophils express the CXC chemokine epithelial cell‐derived neutrophil activating peptide (ENA‐78/CXCL5), a chemokine that can activate neutrophils and in addition possesses angiogenic properties.
Immunocytochemistry detected CXCL5 in eosinophils and the peptide was localized in the specific granules by immunoelectron microscopy.
Methods and Results In eosinophil lysates, 12 ± 2 pg (mean ± SEM) of CXCL5 was detected per 106 cells by enzyme‐linked immunosorbent assay (ELISA). Weak constitutive expression of CXCL5, as well as the related CXC chemokine IL−8/CXCL8, could be detected in freshly isolated eosinophils by RT–PCR. However, during prolonged incubation of eosinophils, a strong increase in both CXCL5 and IL‐8/CXCL8 expression was seen, as detected by RT‐PCR, and increasing amounts of CXCL5 peptide with time were detected in the incubation medium by ELISA. Addition of TNF‐α neutralizing antibodies during prolonged incubation significantly inhibited CXCL5 production, demonstrating involvement of auto‐ and paracrine effects from TNF‐α produced by eosinophils themselves. Addition of IFN‐γ showed a strong inhibitory effect on CXCL5 synthesis.
Conclusion These findings suggest that, through expression of CXCL5, eosinophils can recruit and activate CXC receptor 2 (CXCR2)‐bearing cells such as neutrophils at sites of inflammation. Eosinophils may also promote connective tissue remodelling through release of this peptide.
CREB3L2, a member of the CREB3 family of transcription factors, spans >120 kbp and is composed of 12 exons. We characterized a widely expressed transcript of CREB3L2 generated by an intronic ...polyadenylation site in intron 4 of the gene. It could be translated to a CREB3L2 variant which is localized both in the nucleus and the endoplasmatic reticulum. The protein retains the N-terminal transactivation domain but lacks the DNA-binding domain, the transmembrane domain and the C-terminal part. Experiments using a GAL4 DNA-binding domain fusion model showed that the transcript is a transactivator but it cannot exert its function through the CRE and ATF6 binding sites and has little effect on the GRP78 promoter. Whether this transcript has a cellular function or is targeted for degradation by nonsense-mediated RNA decay system of RNA surveillance is currently unknown.
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