The prognosis of non-small cell lung cancer (NSCLC) is poor, since it has often metastasized to distant organs by the time of diagnosis. Therefore, biomarkers predicting metastasis are crucial. miRNAs play important roles in the regulation of different tumor cell processes, including metastasis. We recently showed that miRNA-214 is linked to a radioresistant phenotype of NSCLC. miRNA-214 has been linked to metastasis in other tumor types. Therefore, we examined the role of miRNA-214 in the metastatic potential of NSCLC. We showed that downregulation of miRNA-214 increased invasive potential, and conversely, overexpression of miRNA-214 decreased invasiveness of NSCLC cells in vitro. Gene expression and bioinformatic analyses of NSCLC cells with ablated miRNA-214, identified a number of metastasis-related target genes, including pregnancy-associated plasma protein A (PAPP-A), alpha protein kinase 2 (ALPK2), cyclin-dependent kinase 6 (CDK6) and tumor necrosis-factor alpha-induced protein 3 (TNFAIP3). These were validated on mRNA and protein level to be regulated by miRNA-214. Through immunoprecipitation we showed that only ALPK2 is directly regulated by miRNA-214. We also examined the protein expression of these four genes in NSCLC tumors with respect to metastatic potential. These results showed that NSCLC tumors express these proteins at moderate-high levels in the nucleus, cytoplasm and/or plasma membrane although with no significant correlation to the overall survival or the metastatic potential of the patients. However, we also showed that the membrane-localized PAPP-A had a higher expression level compared to the cytoplasm-localized. In conclusion, we show that low miRNA-214 expression is linked to a higher invasive potential of NSCLC cells.

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Background Definite noninvasive characterisation of renal tumours positive on 99m Tc-sestamibi single photon emission computed tomography/computed tomography (SPECT/CT) examination including renal oncocytomas (ROs), hybrid oncocytic chromophobe tumours (HOCTs), and chromophobe renal cell carcinoma (chRCC) is currently not feasible. Objective To investigate whether combined 99m Tc-sestamibi SPECT/CT and in situ metabolomic profiling can accurately characterise renal tumours exhibiting 99m Tc-sestamibi uptake. Design, setting, and participants A tissue microarray analysis of 33 tumour samples from 28 patients was used to investigate whether their in situ metabolomic status correlates with their features on 99m Tc-sestamibi SPECT/CT examination. In order to validate emerging data, an independent cohort comprising 117 tumours was subjected to matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI MSI). Outcome measurements and statistical analysis MALDI MSI data analysis and image generation were facilitated by FlexImaging v. 4.2, while k-means analysis by SCiLS Lab software followed by R-package CARRoT analysis was used for assessing the highest predictive power in the differential of RO versus chRCC. Heatmap-based clustering, sparse partial least-squares discriminant analysis, and volcano plots were created with MetaboAnalyst 3.0. Results and limitations We identified a discriminatory metabolomic signature for 99m Tc-sestamibi SPECT/CT–positive Birt-Hogg-Dubè–associated HOCTs versus other renal oncocytic tumours. Metabolomic differences were also evident between 99m Tc-sestamibi–positive and 99m Tc-sestamibi–negative chRCCs, prompting additional expert review; two of three 99m Tc-sestamibi–positive chRCCs were reclassified as low-grade oncocytic tumours (LOTs). Differences were identified between distal-derived tumours from those of proximal tubule origin, including differences between ROs and chRCCs. Conclusions The current study expands the spectrum of 99m Tc-sestamibi SPECT/CT–positive renal tumours, encompassing ROs, HOCTs, LOTs, and chRCCs, and supports the feasibility of in situ metabolomic profiling in the diagnostics and classification of renal tumours. Patient summary For preoperative evaluation of solid renal tumours, 99m Tc-sestamibi single photon emission computed tomography/computed tomography (SPECT/CT) is a novel examination method. To increase diagnostic accuracy, we propose that 99m Tc-sestamibi–positive renal tumours should be biopsied and followed by a combined histometabolomic analysis.

Background: 99mTc-Sestamibi Single Photon Emission Computed Tomography/Computed Tomography (SPECT/CT) contributes to the non-invasive differentiation of renal oncocytoma (RO) from renal cell carcinoma (RCC). We investigated whether standard uptake value (SUV) SPECT, has a beneficial role in differentiating renal oncocytoma (RO) from renal cell carcinoma (RCC) besides visual assessment. As a secondary aim, we evaluated the mitochondrial content of 19 oncocytic tumours arranged in a tissue microarray, by immunohistochemistry, using succinate dehydrogenase complex subunit B (SDHB) protein expression. In addition to visual evaluation of 99mTc-Sestamibi uptake by characterizing renal tumours as Sestamibi positive or Sestamibi negative regarding their uptake compared to the non-tumoral renal parenchyma, SUVmean and SUVmax measurements were performed in the renal tumour and the non-tumoral renal parenchyma. Intra Class Correlation calculated to assess the intra-reader reliability of SUV measurements. ROC-analysis demonstrated an optimal cut off SUV value that differentiates RO from RCC. SDHB score was analysed using Cochran–Armitage test for trend.Results: 57 renal tumours from 52 patients were evaluated. Visual evaluation of 99mTc-Sestamibi SPECT/CT examination resulted in a sensitivity of 83%, whereas quantitative evaluation showed a sensitivity of 64% regarding the differential of RO from RCC. A significant trend (p=0,0328) of increased SDHB score found in the Sestamibi positive group.Conclusion: Quantitative evaluation with SUV SPECT measurements did not improve the performance of 99mTc-Sestamibi SPECT/CT in differentiating RO from RCC. 99mTc- Sestamibi SPECT/CT identified a larger Sestamibi-positive tumour group containing RO, Hybrid Oncocytic Chromophobe Tumours and the majority of chromophobe RCCs. Thus, Sestamibi-negative renal tumours, that are possibly malignant, should be considered for surgery. Patients with Sestamibi-positive tumours can be suited for biopsy and follow up according to active surveillance protocols. Low Grade Oncoytic Tumour, a provisional renal entity with no proven recurrence or metastatic potential, appears to be positive on 99mTc- Sestamibi SPECT/CT examination.