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The spread of tumor cells and the formation of distant metastasis remain the main causes of mortality in cancer patients. However, the mechanisms governing the release of cells from micro-environmental constraints remain unclear. E-cadherin negatively controls the invasion of epithelial cells by maintaining cell–cell contacts. Furthermore, the inactivation of E-cadherin triggers invasion in vitro. However, the role of E-cadherin is complex, as metastasizing cells maintain E-cadherin expression, which appears to have a positive role in the survival of tumor cells. In this report, we present a novel mechanism delineating how E-cadherin function is modulated to promote invasion. We have previously shown that E-cadherin is associated with p100AmotL2, which is required for radial actin formation and the transmission of mechanical force. Here, we present evidence that p60AmotL2, which is expressed in invading tumor cells, binds to the p100AmotL2 isoform and uncouples the mechanical constraint of radial actin filaments. We show for the first time that the coupling of E-cadherin to the actin cytoskeleton via p100AmotL2 is directly connected to the nuclear membrane. The expression of p60AmotL2 inactivates this connection and alters the properties of the nuclear lamina, potentiating the invasion of cells into micropores of the extracellular matrix. In summary, we propose that the balance of the two AmotL2 isoforms is important in the modulation of E-cadherin function and that an imbalance of this axis promotes ameboid cell invasion.

Recent studies suggest that knowledge of differential expression of microRNAs (miRNA) in cancer may have substantial diagnostic and prognostic value. Here, we use a direct sequencing method to characterize the profiles of miRNAs and other small RNA segments for six human cervical carcinoma cell lines and five normal cervical samples. Of 166 miRNAs expressed in normal cervix and cancer cell lines, we observed significant expression variation of six miRNAs between the two groups. To further show the biological relevance of our findings, we examined the expression level of two significantly varying miRNAs in a panel of 29 matched pairs of human cervical cancer and normal cervical samples. Reduced expression of miR-143 and increased expression of miR-21 were reproducibly displayed in cancer samples, suggesting the potential value of these miRNAs as tumor markers. In addition to the known miRNAs, we found a number of novel miRNAs and an additional set of small RNAs that do not meet miRNA criteria. [Cancer Res 2007;67(13):6031–43]

Supplementary Table 6 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Figure 1 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Table 2 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Figure Legends 1-2 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Table 3 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Table 1 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Table 7 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Figure 2 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Table 4 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Supplementary Figure 1 from CREB3L2-PPARγ Fusion Mutation Identifies a Thyroid Signaling Pathway Regulated by Intramembrane Proteolysis

Supplementary Table 5 from Patterns of Known and Novel Small RNAs in Human Cervical Cancer

Building a blastema from the stump is a key step of salamander limb regeneration. Stump-derived cells temporarily suspend their identity as they contribute to the blastema by a process generally referred to as dedifferentiation. Here, we provide evidence for a mechanism that involves an active inhibition of protein syn-thesis during blastema formation and growth. Relieving this inhibition results in a higher number of cycling cells and enhances the pace of limb regeneration. By small RNA profiling and fate mapping of skeletal muscle progeny as a cellular model for dedifferentiation, we find that the downregulation of miR-10b-5p is critical for rebooting the translation machinery. miR-10b-5p targets ribosomal mRNAs, and its artificial upregulation causes decreased blastema cell proliferation, reduction in transcripts that encode ribosomal subunits, dimin-ished nascent protein synthesis, and retardation of limb regeneration. Taken together, our data identify a link between miRNA regulation, ribosome biogenesis, and protein synthesis during newt limb regeneration.

The ATP-regulated K+ channel (KATP) plays an essential role in the control of many physiological processes, and contains a ATP-binding site. Tyrosine kinase inhibitors (TKI) are commonly used drugs, that primarily target ATP-binding sites in tyrosine kinases. Herein, we used the patch-clamp technique to examine the effects of three clinically established TKIs on KATP channel activity in isolated membrane patches, using a pancreatic β-cell line as a KATP channel source. In excised inside-out patches, the activity of the KATP channel was dose-dependently inhibited by imatinib with half-maximal concentration of approximately 9.4 μM. The blocking effect of imatinib was slow and reversible. No effect of imatinib was observed on either the large (KBK) or the small (KSK) conductance, Ca2+-regulated K+ channel. In the presence of ATP/ADP (ratio 1) addition of imatinib increased channel activity approximately 1.5-fold. Sunitinib and nilotinib were also found to decrease KATP channel activity. These findings are compatible with the view that TKIs, designed to interact at the ATP-binding pocket on the tyrosine receptor, also interact at the ATP-binding site on the KATP channel. Possibly, this might explain some of the side effects seen with TKIs.

Abnormalities of the insulin‑like growth factor 2

The Special Issue in Cancers, “The Biological and Clinical Aspects of Merkel Cell Carcinoma”, walks the avid reader through the interesting and sometimes even mysterious facets of Merkel cell carcinoma (MCC), starting at its carcinogenesis to also cover innovative treatment options [...]

The ATP-regulated K

Merkel cell carcinoma (MCC) is a rare and aggressive skin cancer where Merkel cell Polyomavirus (MCPyV) contributes to the pathogenesis. In an adjuvant setting, radiotherapy (RT) is believed to give a survival benefit. The prognostic impact of sex related to MCPyV-status and adjuvant RT were analyzed in patients referred to Karolinska University Hospital. Data were collected from 113 patients&rsquo, hospital records and MCPyV analyses were made in 54 patients (48%). We found a significantly better overall survival (OS) for women compared to men and a significant difference in OS in patients receiving adjuvant RT. Furthermore, we found that men with virus negative MCC have an increased risk for earlier death (HR 3.6). This indicates that MCPyV positive and negative MCC act as two different diseases, and it might be due to different mechanism in the immune response between male and female patients. This could have significance in tailoring treatment and follow-up in MCC patients in the future.

Essentials It is unclear if platelet micro-RNAs can regulate de novo protein synthesis of platelets. Platelet de novo protein synthesis of thrombospondin-1 (TSP-1) was induced by thrombin. Thrombin stimulation in vitro altered platelet microRNA profiles, including decreased miR-27b. Decreased miR-27b hampers platelet angiogenic activities via enhancing de novo TSP-1 synthesis. Summary Background Platelets can synthesize proteins upon activation. Platelets contain a number of microRNAs (miRNA) and a fully functional miRNA effector machinery. It is, however, unclear if platelet miRNAs can regulate protein synthesis of platelets, and whether the regulation may produce a physiological impact. Objectives To investigate if and how platelet miRNAs regulate de novo syntheses of angiogenic regulators and subsequently modulate platelet angiogenic activities. Methods and Results Microarray-based miRNA profiling showed that thrombin stimulation in vitro down- or up-regulated a number of platelet miRNAs, both in the total platelet miRNAs and in Ago2-associated miRNAs. Among those altered miRNAs, miR-27b was down-regulated in both the total and Ago2-immunoprecipitated miRNA profiles of platelets, which was confirmed by reverse transcription-quantitative PCR (RT-qPCR). Using western blotting assays, we showed that thrombin induced platelet de novo synthesis of thrombospondin-1, and that the level of thrombospondin-1 synthesis could reach a level of 3-5-fold higher than that before thrombin stimulation. With either the platelet precursor megakaryocyte cell line MEG-01 cells or mature platelets, we demonstrated that transfection of miR-27b mimic, but not the negative control of miRNA mimic, markedly reduced thrombospondin-1 protein levels. The latter subsequently enhanced platelet-dependent endothelial tube formation on matrigel. Conclusions Thrombin stimulation in vitro reduces platelet miR-27b levels that may markedly enhance thrombin-evoked platelet de novo synthesis of thrombospondin-1. Elevation of platelet miR-27b by transfection inhibits thrombospondin-1 synthesis, and subsequently enhances platelet pro-angiogenic activities. Hence, platelet activation-dependent reduction of miR-27b levels may represent a novel negative regulatory mechanism of platelet angiogenic activities.

Ovarian germ cell tumors (OGCTs) and sex cord stromal tumors (SCSTs) are rare gynecologic tumors that are derived from germ and stromal cells, respectively. Unlike their epithelial counterparts, molecular pathogenesis of these tumor types is still poorly understood. Here, we characterized microRNA (miRNA) expression profiles of 9 OGCTs (2 malignant and 7 benign) and 3 SCSTs using small RNA sequencing. We observed significant miRNA expression variations among the three tumor groups. To further demonstrate the biological relevance of our findings, we selected 12 miRNAs for validation in an extended cohort of 16 OGCTs (9 benign and 7 malignant) and 7 SCSTs by reverse transcription-quantitative polymerase chain reaction. Higher expression of miR‑373‑3p, miR‑372‑3p and miR‑302c‑3p and lower expression of miR‑199a‑5p, miR‑214‑5p and miR‑202‑3p were reproducibly observed in malignant OGCTs as compared to benign OGCTs or SCSTs. Comparing with benign OGCTs, miR‑202c‑3p and miR‑513c‑5p were more abundant in SCSTs. Additionally, we examined Beclin 1 (BECN1), a target of miR‑199a‑5p, in the clinical samples using western blot analysis. Our results show that BECN1 expression was higher in malignant OGCTs than benign OGCTs, which is concordant with their lower miR‑199a‑5p expression. This study suggests that these miRNAs may have potential value as tumor markers and implications for further understanding the molecular basis of these tumor types.

High-risk human papillomavirus (HPV) testing is a recommended triage approach for females with atypical squamous cells of undetermined significance (ASCUS), but due to its poor specificity this approach is not recommended for patients with low-grade squamous intraepithelial lesions (LSIL). The objective of the current study was to determine microRNA (miR)-205 expression levels in liquid-based cytology (LBC) samples, and evaluate their ability to predict cervical intraepithelial neoplasia grade 2/3 or worse (CIN2/3+) in females with minor cytological abnormalities. LBC samples were obtained from patients attending the Swedish Cervical Cancer Screening Program. The Mann-Whitney U test, one-way analysis of variance, Kruskal-Wallis test, Spearman rank order correlation analysis, and Pearson's χ2 test were used to assess the results. Accuracy analyses indicated that high miR-205 expression had a significantly higher specificity to high-risk HPV testing, and a sensitivity similar to that of high-risk HPV testing to predict CIN2+ and CIN3+ in women with LSIL, but not those with high-grade squamous intraepithelial lesions. Although further research is required for females with LSIL, miR-205 expression in LBC samples may be a novel triage marker for, or a beneficial supplement to high-risk-HPV testing in these patients.

miR-223-3p is deregulated in several tumor types and plays an important role in tumorigenesis and progression. However, its role in the pathogenesis of testicular germ cell tumor (TGCT) remains uncharacterized. We previously demonstrated that miR-223-3p expression was increased in TGCTs compared with normal testes (NT), suggesting that miR-223-3p may have an oncogenic role in TGCT. Using published dataset and The Cancer Genome Atlas database, we validated higher miR-223-3p expression in TGCTs than NT, and found a negative correlation between miR-223-3p and FBXW7 mRNA expression levels. Using both gain- and loss-of-function experiments, we show that miR-223-3p regulates FBXW7 protein expression, cell growth and apoptosis in TGCT cell lines. Additionally, we demonstrate that ectopic expression of the full-length coding sequence of FBXW7 could rescue the cell growth and apoptotic effects mediated by miR-223-3p. Our findings suggest an oncogenic role for miR-223-3p in TGCT, which promotes cell growth and inhibits apoptosis through repression of FBXW7.

Viruses can inhibit host autophagy through multiple mechanisms, and evasion of autophagy plays an important role in immune suppression and viral oncogenesis. Merkel cell polyomavirus (MCPyV) T‐antigens are expressed and involved in the pathogenesis of a large proportion of Merkel cell carcinoma (MCC). Yet, how MCPyV induces tumorigenesis is not fully understood. Herein, we show that MCPyV T‐antigens induce miR‐375, miR‐30a‐3p and miR‐30a‐5p expressions, which target multiple key genes involved in autophagy, including ATG7, SQSTM1 (p62) and BECN1. In MCC tumors, low expression of ATG7 and p62 are associated with MCPyV‐positive tumors. Ectopic expression of MCPyV small T‐antigen and truncated large T‐antigen (LT), but not the wild‐type LT, resulted in autophagy suppression, suggesting the importance of autophagy evasion in MCPyV‐mediated tumorigenesis. Torin‐1 treatment induced cell death, which was attenuated by autophagy inhibitor, but not pan‐caspase inhibitor, suggesting a potential role of autophagy in promoting cell death in MCC. Conceptually, our study shows that MCPyV oncoproteins suppress autophagy to protect cancer cells from cell death, which contribute to a better understanding of MCPyV‐mediated tumorigenesis and potential MCC treatment., What's new? About four‐fifths of Merkel cell carcinomas harbor the Merkel cell polyomavirus (MCPyV) genome, though mutations in viral T antigens generally render MCPyV replication‐deficient. Here, the authors describe a network by which MCPyV oncoproteins and viral‐regulated miRNAs hijack autophagy machinery in MCC. In cells, expression of MCPyV T‐antigens induced miR‐375, miR‐30a‐3p, and miR‐30a‐5p expression. Subsequent experiments validated these miRNAs as autophagy regulators that specifically target the genes ATG7, SQSTM1, and BECN1. Autophagy suppression protected MCC cell survival. This novel insight into the role of MCPyV and autophagy regulation in MCC may be relevant in the generation of future therapeutic strategies.

MicroRNA-375 (miR-375) is deregulated in multiple tumor types and regulates important targets involved in tumorigenesis and metastasis. This miRNA is highly expressed in Merkel cell carcinoma (MCC) compared to normal skin and other non-MCC skin cancers, and its expression is high in Merkel cell polyomavirus (MCPyV)-positive (MCPyV+) and low in MCPyV-negative (MCPyV&minus, ) MCC tumors. In this study, we characterized the function and target of miR-375 in MCPyV+ and MCPyV&minus, MCC cell lines. Ectopic expression of miR-375 in MCPyV&minus, MCC cells resulted in decreased cell proliferation and migration, as well as increased cell apoptosis and cell cycle arrest. However, in MCPyV+ MCC cells, inhibition of miR-375 expression reduced cell growth and induced apoptosis. Additionally, the expression of lactate dehydrogenase B (LDHB), a known target of miR-375, was inversely correlated with miR-375. Silencing of LDHB reduced cell growth in MCPyV&minus, cell lines, while its silencing in MCPyV+ cell lines rescued the cell growth effect mediated by miR-375 inhibition. Together, our results suggest dual roles of miR-375 and LDHB in MCPyV and non-MCPyV-associated MCCs. We propose that LDHB could be a therapeutic target in MCC and different strategies should be applied in virus- and non-virus-associated MCCs.

The use of imatinib mesylate has greatly improved the clinical outcome for gastrointestinal stromal tumor (GIST) patients. However, imatinib resistance is still a major clinical challenge, and the molecular mechanisms are not fully understood. We have previously shown that miR-125a-5p and its mRNA target PTPN18 modulate imatinib response in GIST cells. Herein, we evaluated phosphorylated FAK (pFAK) as a candidate downstream target of PTPN18 and the possible association of this regulation with imatinib resistance in GIST. FAK and pFAK expressions were evaluated in GIST882 cells transfected with short hairpin RNA or short interfering RNA targeting PTPN18 or miR-125a-5p mimic, imatinib-resistant GIST882R subclones and clinical samples using Western blot analyses. FAK phosphorylation was blocked using the FAK inhibitor 14 (FAKi) and the effects on cell viability and apoptosis were evaluated using WST-1 assay and cleaved PARP expression. Clinical associations of FAK and pFAK expression with imatinib resistance, KIT mutation and patient outcome were assessed by Fisher's exact test or log-rank test. Over-expression of miR-125a-5p and silencing of PTPN18 increased pFAK, but not FAK, expression in GIST cells. Higher pFAK expression was observed in the GIST882R subclones with acquired imatinib resistance compared to their imatinib-sensitive parental cells. Treatment with FAKi in imatinib-resistant GIST882R cells reduced cell viability and increased apoptosis upon imatinib treatment. Additionally, FAKi could rescue the imatinib resistance effect mediated by miR-125a-5p over-expression. In clinical samples, high FAK and pFAK expressions were associated with KIT mutation status, and high FAK expression was also associated with metastasis in GIST. Higher pFAK was found in cases with shorter overall survival. Our findings highlight an important role for miR-125a-5p regulation and its downstream target pFAK for imatinib resistance in GIST. pFAK and FAK may have prognostic values in GIST.

Strategies for correct diagnosis, treatment evaluation and recurrence prediction are important for the prognosis and mortality rates among cancer patients. In spite of major improvements in clinical management, gastrointestinal stromal tumors (GISTs) can still be deadly due to metastasis and recurrences, which confirms the unmet need of reliable follow-up modalities. Tumor-specific secreted, shed or leaked proteins (collectively known as secretome) are considered promising sources for biomarkers, and suitable for detection in biofluids. Herein, we stimulated cell secretion in the imatinib-sensitive GIST882 cell line and profiled the secretome, collected as conditioned media, by using a shotgun proteomics approach. We identified 764 proteins from all conditions combined, 51.3% being predicted as classically/non-classically secreted. The protein subsets found were dependent on the stimulatory condition. The significant increase in protein release by the classical pathway was strongly associated with markers already found in other cancer types. Furthermore, most of the released proteins were non-classically released and overlapped to a high degree with proteins of exosomal origin. Imatinib pre-treatment radically changed these secretory patterns, which can have clinical implications when investigating biomarkers in imatinib-treated versus non-treated GIST patients. Our results show, for the first time, that GISTs contain a secretome signature. In the search for suitable biomarkers in the more complex GIST patient samples, this study aids in the understanding of basic GIST secretome characteristics.

Human cytomegalovirus (HCMV) infection results in the production of virions, dense bodies (DBs) and non-infectious enveloped particles, all of which incorporate proteins and RNAs that can be transferred to host cells. Here, we investigated whether virions and DBs also carry microRNAs (miRNAs) and assessed their delivery and functionality in cells. Human lung fibroblasts (MRC-5) were infected with the HCMV strain AD169, and conditioned cell culture medium was collected and centrifuged. The pellets were treated with RNase-ONE, and the virions and DBs were purified with a potassium tartrate–glycerol gradient and dialysed. The virions and DBs were incubated with micrococcal nuclease, DNA and RNA were extracted and then analysed with TaqMan PCR assays, while the proteins were examined with Western blots. To assess the delivery of miRNAs to cells and their functionality, virions and DBs were irradiated with UV light. The purity of the virions and DBs was confirmed by typical morphology, the presence of the structural protein pp65 and the HCMV genome, the ability to infect MRC-5 cells and the absence of the host genome. RNA analysis revealed the presence of 14 HCMV-encoded miRNAs (UL22A-5p, US25-1-5p, UL22A-3p, US5-2-3p, UL112-3p, US25-2-3p, US25-2-5p, US33-3p, US5-1, UL36-5p, US4-5p, UL36-3p, UL70-5p and US25-1-3p), HCMV immediate-early mRNA and long non-coding RNA2.7, moreover, two host-encoded miRNAs (hsa-miR-218-5p and hsa-miR-21-5p) and beta-2-microglobulin RNA. UV-irradiated virions and DBs delivered viral miRNAs (US25-1-5p and UL112-3p) to the host cells, and miR-US25-1-5p was functional in a luciferase reporter assay. We conclude that virions and DBs carry miRNAs that are biologically functional and can be delivered to cells, which may affect cellular processes.

e21047 Background: Merkel cell carcinoma (MCC) is a rare and aggressive neuroendocrine skin cancer. The pathogenesis is linked to the immune system and Merkel cell polyoma virus (MCPyV) is present in about 80% of MCC. Reports have shown that patients with a virus-negative MCC have a worse prognosis. Adjuvant radiotherapy given after surgery has previously been shown to give a survival benefit in retrospective data. Our aim is to analyze clinical variables and their prognostic impact in a Swedish cohort. Methods: All patients with confirmed diagnosis of MCC referred to Karolinska University Hospital in Sweden from 1989-2018 were included and retrospective data was collected. Results: 113 patients were included. Median age at operation was 75.4 years (range 19-99); 64 (57%) were female. Treatment included surgery with (n = 46) or without adjuvant radiotherapy (RT). 43% of female patients and 37% of male patients received RT. Median overall survival (OS) was 2,6 years (range 0,1-25,8), for female patients 3,4 years (range 0,1-25,8) and for male patients 1,8 years (range 0,1-19,0). Median OS in surgery group was 1,8 years (range 0,4-25,8). With adjuvant RT median OS improved with 1,7 years (p = 0,0001) to 3,5 years (range 0,2-22,1), for female patients 4,0 years (range 0,2-22,1) and for male patients 2,0 years (range 0,3-8,8). MCPyV status was available in 53 patients. 74% were MCPyV positive, 26% negative. MCPyV gave no difference in OS, however when gender was added in to multivariate analysis, female patients with virus negative disease had better outcome than virus negative males (p = 0.03). In virus positive MCC there was no difference in OS between female and male patients. Conclusions: Our data confirms the positive impact of adjuvant radiotherapy on survival and illustrates the difference in outcome between female and male patients, with women having a better outcome. In the MCPyV group the tendency is that men with virus negative MCC has the worst outcome. Our findings indicate that MCPyV positive and negative MCC act as two different diseases and it also raises questions if there is a difference in the disease itself or the immune response towards MCC between male and female patients.

Introduction Background and aim Merkel cell carcinoma (MCC) is an aggressive type of skin cancer. About 80% of MCCs harbour integrated Merkel cell polyomavirus (MCV) genome with a mutation in the large T (LT) gene. MCV T-antigens are required for neoplastic transformation and maintenance of cell growth. however the molecular mechanism by which the virus induces tumorigenesis remains unclear. In this study we aimed to identify and characterise functional role of MCV-regulated microRNAs in MCC. Material and methods We did silencing or ectopic expression of MCV T-antigens to identify specific miRNAs regulated by MCV T-antigens by RT-qPCR. The involvement of MCV T-antigens and miRNAs in autophagy was evaluated using LC3-II conversion, mRFP-EGFP-LC3 reporter and/or transmission electron microscopy. The targets of miRNAs were verified by western blot analysis and luciferase reporter assays. We also look for the DICER expression by western blot. Results and discussions We identified specific miRNAs regulated by MCV T-antigens. Using both gain- and loss-of-function experiments, we showed that MCV T-antigens and MCV-regulated miRNAs (miR-375, miR-30a-5p and miR-30a-3p) could regulate autophagy. We further demonstrated that miR-375 could directly regulate ATG7 and SQSTM1, while both miR-30a-3p and miR-30a-5p could target BECN1. We also identified the MCV T-antigens regulates these miRNAs through post-transcriptionally by regulating DICER expression in MCV positive MCC. Conclusion We provide evidence that MCV T-antigens regulate miRNAs and autophagy in MCC. MCV T-antigens increase miRNA expressions through pots-transcriptional regulation, and several of these miRNAs can directly target multiple autophagy genes that lead to suppression of autophagy in MCC.

e21601Background: Merkel cell carcinoma (MCC) is a rare and aggressive neuroendocrine skin cancer. The pathogenesis is linked to the immune system and immunocompromised patients have an increased r...

INTRODUCTION:MicroRNAs (miRNAs) are small non-coding RNA that plays a vital role in cancer progression. Neo-adjuvant chemotherapy (NAC) has become the standard of care for locally advanced breast cancer. The aim of this study was to evaluate miRNA alterations during NAC using multiple samples of tissue and serum to correlate miRNA expression with clinico-pathological features and patient outcomes. METHODS:Tissue and serum samples were collected from patients with locally advanced breast cancer undergoing NAC at four time points: time of diagnosis, after the first and fourth cycle of doxorubicin/cyclophosphamide treatment, and after the fourth cycle of docetaxel administration. First, we evaluated the miRNA expression profiles in tissue and correlated expression with clinico-pathological features. Then, a panel of four miRNAs (miR-451, miR-3200, miR-21, and miR-205) in serum samples was further validated using quantitative reverse-transcription polymerase chain reaction (RT-qPCR). The alterations in serum levels of miRNA, associations with clinical and pathological responses, correlation with clinico-pathological features, and survival outcomes were studied using Friedman, Mann-Whitney U, and Spearman, Wilcoxon signed-ranks tests. P≤0.05 was considered statistically significant. RESULTS:We analyzed 72 tissue samples and 108 serum samples from 9 patients and 27 patients, respectively. MicroRNA expression profiling of tumor versus normal tissue revealed more than 100 differentially expressed miRNAs. Serum miR-451 levels were significantly decreased during treatment, and higher serum levels were associated with improved clinical and pathological responses and disease-free survival. This is one of the early reports on miR-3200 in response to treatment in breast cancer, as serum levels of miR-3200 found to decline during NAC, and higher serum levels were associated with lower residual breast cancer burden and relapse rates at time of diagnosis. CONCLUSION:Variations in serum miRNA levels during NAC treatment may be therapeutically significant for predicting response and survival outcomes.

Gastrointestinal stromal tumor (GIST) is the most commonly diagnosed mesenchymal tumor in the gastrointestinal tract. This tumor type is driven by gain-of-function mutations in receptor tyrosine kinases (such as KIT, PDGFRA, and BRAF) or loss-of-function mutations in succinate dehydrogenase complex subunit genes (SDHx). Molecular studies on GIST have improved our understanding of the biology of the disease and have led to the use of targeted therapy approach, such as imatinib for KIT/PDGFRA-mutated GIST. Recently, microRNAs have emerged as important regulators of KIT expression, cancer cell behavior, and imatinib response in GIST. This chapter aims to provide an overview on current understanding of the biological roles of microRNAs in GIST and possible implications in prognosis and therapeutic response.

Deregulation of microRNAs (miRNAs) contributes to the development and progression of many cancer types; however, their functions in the pathogenesis of testicular germ cell tumor (TGCT) remain unclear. Here, we determined miRNA expression profiles of TGCTs and normal testes using small RNA sequencing, and identified several deregulated miRNAs in TGCTs, including the miR-506~514 cluster. In functional studies in vitro we demonstrated that miR-514a-3p induced apoptosis through direct regulation of the paternally expressed gene 3 (PEG3), and ectopically expressed PEG3 could rescue the apoptotic effect of miR-514a-3p overexpression. Silencing of PEG3 or miR-514a-3p overexpression reduced nuclear accumulation of p50 and NF-κB reporter activity. Furthermore, PEG3 was co-immunoprecipitated with tumor necrosis factor receptor-associated factor 2 (TRAF2) in TGCT cell lysates. We propose a model of PEG3-mediated activation of NF-κB in TGCT. Loss of miR-514a-3p expression in TGCT increases PEG3 expression that recruits TRAF2 and activates the NF-kappa B pathway, which protects germ cells from apoptosis. Importantly, we observed strong expression of PEG3 and nuclear p50 in the majority of TGCTs (83% and 78%, respectively). In conclusion, our study describes a novel function for miR-514a-3p in TGCT and highlights an unrecognized mechanism of PEG3 regulation and NF-κB activation in TGCT.

s / Journal of Clinical Virology 70 (2015) S1–S126 S117 confirmed with recomLine imunoblot test. Corresponding values for Dia.Pro test were 2.5% and 1,1%, respectively. This work is supported by IGA grant NT/13884/2012 of Ministry of Health of the Czech Republic http://dx.doi.org/10.1016/j.jcv.2015.07.270