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Background Total knee arthroplasty surgery (TKA) using prenavigated Patient Specific Instruments (PSI) technique represents one of the most recent technological evolutions in development of prosthetic surgery. The aim of this study was to evaluate kinematic and functional recovery of patients operated with prenavigated PSI technique compared to those operated with traditional technique. Methods A cohort of 20 patients is divided in two groups; some are operated with traditional technique (with NexGen Knee system) and others with prenavigated PSI technique (with eMP Knee system) at Asiago Hospital. Limb circumferences are measured for edema evaluation and different evaluation forms are provided to patients: SF-36, KSS pre-surgery (T0), KSS 15 (T1) and 45 days after surgery (T2). Gait Analysis is performed 60 days post-surgery, after leaving crutches. Results The analysis of KSS and SF-36 evaluation forms shows a greater improvement in PSI Evolution group in terms of articulation (comparison between T0 and T1), knee function and early return to physical and social activities. Pain is lesser in NexGen group, in an earlier phase, but 45 days after surgery (T2) there are no significant differences between two groups. Perception of general state of health improves more and earlier in NexGen. In NexGen group edema evaluation had significant differences at the level of prosthetic leg, but not in knee and thigh. Overall: the walking pattern is more physiological in PSI Evolution group. Conclusions The present study highlighted the superiority of prenavigated PSI technique over traditional technique in recovering functionality of prosthetic knee and in restoring a more physiological path pattern.

Background Breast metastases from colorectal cancer (CRC) are very uncommon. There is no unanimous consensus regarding the best treatment for this rare condition, and management is, especially in elderly patients, limited to diagnosis and palliative care. Capecitabine, an oral fluoropyrimidine derivative, might be helpful in controlling the disease and may be a treatment option for patients unable to receive more aggressive chemotherapy. Case summary We report a case of synchronous massive breast metastasis from CRC in an 85 year old patient who came to the hospital presenting a huge mass originating from the axillary extension of the right breast. A whole body computed tomography also showed a mass in the right colon. The patient underwent a simple right mastectomy along with right hemicolectomy. The resected breast showed massive metastasis from CRC with intense and homogeneous nuclear CDX2 staining, while the colon specimen revealed poorly differentiated adenocarcinoma stage pT4a pN0 pM1 (breast) (Tumor Node Metastasis 2017). Three months later she developed a subcutaneous mass at the site of the previous mastectomy. An ultrasound guided biopsy was carried out again and revealed a metastasis from CRC. The patient then started treatment with capecitabine plus bevacizumab, obtaining stable disease (RECIST criteria) and a clinical benefit after 3 mo of therapy. Conclusion In our experience, capecitabine and bevacizumab may be a useful treatment option for breast metastases from primary CRC in elderly patients.

A simple and efficient method using the fireworks algorithm (FWA) is presented for pruning the search space of an impulsive maneuver transfer from a user-defined low Earth orbit (LEO) to a desired ...

This paper focuses on the trajectory design for missions destined to explore Mars and/or Phobos departing from Low Earth Orbit (LEO) and arriving into a Mars-Phobos Distant Retrograde Orbit (DRO). Lunar DROs are also briefly explored as an alternative departure location. A Mars-Phobos DRO is a relatively stable environment which would make both the surfaces of Mars and Phobos available for a reasonable propellant expenditure. This paper presents the methodology used to compute LEO to Mars-Phobos DRO trajectories and results regarding required C 3 at launch, v ∞ at arrival, Time-of-Flight ( T O F ), and total Δ V for various Mars-Phobos DROs using full ephemeris planetary data. The results show that propellant-optimal trajectories from LEO to a specified Mars-Phobos DRO could be used as a staging location between Mars and Phobos. Assuming that refueling is available at the targeted DRO, LEO to Low Mars Orbits (LMO) trajectories would have higher total Δ V due to the additional stop at the Mars-Phobos DRO. However, the aformentioned trajectories would have lower Initial Mass in LEO (IMLEO) and thus a lower gear ratio thanks to the added “pit stop” located at the given DRO. This results in a lower overall spacecraft dry mass that needs to be launched into space from Earth's surface.

Background Respiratory complications caused by the inability to protect the upper airways and ineffective cough represent a major cause of morbidity and mortality in patients with cerebral palsy (CP). Even though the application of positive end-expiratory pressure (PEP) through a face mask has gained large popularity as a technique to prevent bronchial mucous encumbrance, its long-term effects on clinical course and respiratory function in individuals with CP have not been investigated. Aim The aim of this study is to investigate whether regular application of PEP through a face mask can improve clinical status and respiratory function in patients with severe CP. Design Observational, retrospective cohort study. Setting The outpatient rehabilitation unit of the IRCCS E. Medea Rehabilitation Hospital in Conegliano, Italy. Population CP outpatients admitted to the unit between January 1st, 2006 and December 31st, 2018. Methods All the medical records of the enrolled patients were collected and reviewed. All patients underwent multidisciplinary respiratory evaluation at T0 (immediately before the beginning of PEP-use) and T1 (12 months after). The evaluation assessed respiratory infections history (number of exacerbations per year), blood gas analysis, measurement of airway resistance through the interrupter technique. Results Twenty-one patients affected with CP (mean age 9.19±5.56 years, range 3-23 years, 8 females) were included. All patients had more than 3 infections per year (mean 4.81±1.17) in the year prior to treatment (T0). At T1 mean number of infections was 1.57±0.81); 17 patients (80%) reported less than three infections; two patients (10%) reported zero infections, two patients (10%) reported three infections. Blood gas analysis and airway resistance values did not show a significant difference at T0 and T1. Conclusions Daily PEP-mask therapy reduces frequency of respiratory exacerbations in patients with severe bilateral CP. Clinical rehabilitation impact PEP-mask is a valuable rehabilitative tool in severe CP patients with frequent respiratory exacerbations.

miRNAs play a central role in the complex signaling network of cancer cells with the tumor microenvironment. Little is known on the origin of circulating miRNAs and their relationship with the tumor microenvironment in lung cancer. Here, we focused on the cellular source and relative contribution of different cell types to circulating miRNAs composing our risk classifier of lung cancer using in vitro/in vivo models and clinical samples. A cell‐type specific expression pattern and topography of several miRNAs such as mir‐145 in fibroblasts, mir‐126 in endothelial cells, mir‐133a in skeletal muscle cells was observed in normal and lung cancer tissues. Granulocytes and platelets are the major contributors of miRNAs release in blood. miRNAs modulation observed in plasma of lung cancer subjects was consistent with de‐regulation of the same miRNAs observed during immunosuppressive conversion of immune cells. In particular, activated neutrophils showed a miRNA profile mirroring that observed in plasma of lung cancer subjects. Interestingly mir‐320a secreted by neutrophils of high‐risk heavy‐smokers promoted an M2‐like protumorigenic phenotype through downregulation of STAT4 when shuttled into macrophages. These findings suggest a multifactorial and nonepithelial cell‐autonomous origin of circulating miRNAs associated with risk of lung cancer and that circulating miRNAs may act in paracrine signaling with causative role in lung carcinogenesis and immunosuppression., What's new? microRNAs play a central role in the complex signaling network of cancer cells with the tumor microenvironment. However, little is known on the origin of circulating miRNAs and their mechanisms of action. This study found a multifactorial and non‐epithelial cell‐autonomous origin of circulating miRNAs associated with lung cancer risk. The findings also suggest a link between an immunosuppressive and pro‐tumorigenic microenvironment and modulation of circulating miRNAs associated with lung cancer risk. The authors propose a novel mechanism whereby miRNA released by neutrophils induce macrophage polarization to support lung cancer growth, highlighting the potential for reprogramming macrophages toward an anti‐tumor polarization.

The development of a minimally invasive test, such as liquid biopsy, for early lung cancer detection in its preclinical phase is crucial to improve the outcome of this deadly disease. MicroRNAs (miRNAs) are tissue specific, small, non-coding RNAs regulating gene expression, which may act as extracellular messengers of biological signals derived from the cross-talk between the tumor and its surrounding microenvironment. They could thus represent ideal candidates for early detection of lung cancer. In this work, a methodological workflow for the prospective validation of a circulating miRNA test using custom made microfluidic cards and quantitative Real-Time PCR in plasma samples of volunteers enrolled in a lung cancer screening trial is proposed. In addition, since the release of hemolysis-related miRNAs and more general technical issues may affect the analysis, the quality control steps included in the standard operating procedures are also presented. The protocol is reproducible and gives reliable quantitative results; however, when using large clinical series, both pre-analytical and analytical features should be cautiously evaluated.

Digital PCR (dPCR) is an innovative approach for detection and quantification of nucleic acid that offers an alternative method to conventional real-time quantitative PCR for absolute quantification. dPCR is a highly precise and sensitive technique that does not require a standard reference, making it a suitable method for the detection of microRNAs. The potential of these small noncoding RNA as biomarkers is on the rise, especially due to their presence in body fluids, making them easily accessible. Nevertheless, the problem of lack of consensus regarding an optimal method for miRNAs normalization compromises their use. Here, we describe an innovative method for the absolute quantification of miRNAs across different types of biological samples using a chip-based platform, the QuantStudio 3D digital PCR.

Digital PCR (dPCR) is an innovative approach for detection and quantification of nucleic acid that offers an alternative method to conventional real-time quantitative PCR for absolute quantification. dPCR is a highly precise and sensitive technique that does not require a standard reference, making it a suitable method for the detection of microRNAs. The potential of these small noncoding RNA as biomarkers is on the rise, especially due to their presence in body fluids, making them easily accessible. Nevertheless, the problem of lack of consensus regarding an optimal method for miRNAs normalization compromises their use. Here, we describe an innovative method for the absolute quantification of miRNAs across different types of biological samples using a chip-based platform, the QuantStudio 3D digital PCR.

Lung cancer is the most common cause of cancer deaths worldwide and numerous ongoing research efforts are directed to identify new strategies for its early detection. The development of non-invasive blood-based biomarkers for cancer detection in its preclinical phases is crucial to improve the outcome of this deadly disease. MicroRNAs (miRNAs) are a new promising class of circulating biomarkers for cancer detection and prognosis definition, but lack of consensus on data normalization methods for circulating miRNAs and the critical issue of haemolysis, has affected the identification of circulating miRNAs with diagnostic potential. We describe here an interesting approach for profiling circulating miRNAs in plasma samples based on the evaluation of reciprocal miRNA levels measured by quantitative Real-Time PCR. By monitoring changes of plasma miRNA-ratios, it is possible to assess the deregulation of tumor-related miRNAs and identify signatures with diagnostic and prognostic value. In addition, to avoid bias due to the release of miRNAs from blood cells, a miRNA-ratios signature distinguishing haemolyzed samples was identified. The method described was validated in plasma samples of lung cancer patients, but given its reproducibility and reliability, could be potentially applied for the identification of diagnostic circulating miRNAs in other diseases.

Uterine leiomyosarcoma (LMS) is a rare tumor constituting 1% of all uterine malignancies. This sarcoma demonstrates an aggressive growth pattern with an high rate of recurrence with hematologic dissemination; the most common sites are lung, liver, and peritoneal cavity, head and neck district being rarely interested. Only other four cases of metastasis in the oral cavity have been previously described. The treatment of choice is surgery and the use of adjuvant chemotherapy and radiation has limited impact on clinical outcome. In case of metastases, surgical excision can be performed considering extent of disease, number and type of distant lesions, disease free interval from the initial diagnosis to the time of metastases, and expected life span. We illustrate a case of uterine LMS metastasis in the upper buccal gingiva that occurred during chemotherapy in a 63-year-old woman that underwent a total abdominal hysterectomy with bilateral salpingo-oophorectomy for a diagnosis of LMS staged as pT2bN0 and that developed lung metastases eight months after primary treatment. Surgical excision of the oral mass (previously misdiagnosed as epulis at a dental center) and contemporary reconstruction with pedicled temporalis muscle flap was performed in order to improve quality of life. Even if resection was achieved in free margins, “local” relapse was observed 5 months after surgery.

This paper presents the conceptual design of the IMaGInE (Innovative Mars Global International Exploration) Mission whose mission objectives are to deliver a crew of four astronauts to the surface of Deimos and a robotic exploration mission to Phobos for approx-imately 343 days during the years 2031 and 2032, perform surface excursions, technology demonstrations, and In Situ Resource Utilization (ISRU) of the Martian moons as well as site reconnaissance for future human exploration of Mars. This is the winning mission design of the 2016 Revolutionary Aerospace Systems Concepts-Academic Linkage (RASC-AL) competition, awarded with the "Best in Theme," "Best Overall," and "Pio-neering Exceptional Achievement Concept Honor (PEACH)" prizes. This competition was sponsored by NIA and NASA.

Lung cancer is the leading cause of cancer-related mortality in the United States and Europe. The median 5-year survival rate for patients with non– small celllung cancer(NSCLC)isabout 15%.Relative survival for lung tumors is strongly dependent from stage at diagnosis: 5-years survival is 49%, 16%, and 2% for patients with early, locally advanced, and metastatic disease, respectively. 1,2 The natural history of lung cancers is influenced by the acquired ability of cancer cells to invade lymphatic and vascular vessels resulting in nodal and distant metastases. In particular, lymphatic spreading generally follows the pulmonary artery branching system: lower lobe lymphatics generally drain to the posterior mediastinum and to the subcarinal nodes; right upper lobe drains to the superior mediastinum; and left upper lobe drains to the anterior and superior mediastinum. The determination of stage at diagnosis is fundamental in terms of prognostic and therapeutic implications. To plan the best therapeutic strategy, the most important clinical features are the detection of distant metastases and, in nonmetastatic disease,theidentificationandlocalization ofpathologic thoracic lymph nodes by station. Histologic type,tumorsizeandlocation,involvementofpleura, tumor grade, performance status, and biological features are the other cornerstones of prognosis for NSCLC. Surgery, chemotherapy, target agents, and radiotherapy are the therapeutic options available to treat NSCLC. Surgery is the only really curative treatment choice, but its outcome is still poor, in particular for patients with mediastinal lymph node involvement. Good long-term survival is obtained in stages I and II after pulmonary resection, whereas patient survival after surgery in stage IIIA-N2 is still disappointing. Data from several clinical trials show that in completely resected early stages, the 5-year recurrence rates for patients in stage I, stage II-N0, and stage II-N1 were 16%, 39%, and 46%, respectively. 3,4 On the other side, patients with surgically treated stage IIIA-N2 NSCLC have a 5-year recurrence rate and a 5year overall survival (OS) rate of 10% to 15%. In particular, patients with bulky mediastinal involvement have a 5-year survival rate of 2% to 5%. 4 High frequency of distant failure after surgical resection of NSCLC with nodal metastasis suggests that nodal invasion could be considered an indicator of systemic metastasis. Multimodal treatments have been investigated to maximize the gain in survival for patients with metastasis in regional lymph nodes or locally advanced disease.

Introduction Genetic alterations suitable for targeted therapy are poorly known issues in pulmonary sarcomatoid carcinoma (PSC), an uncommon and life-threatening family of non–small cell lung cancers. Methods Ninety-eight PSCs were assessed for MNNG HOS Transforming gene ( MET ) and anaplastic lymphoma receptor tyrosine kinase gene ( ALK ) status by fluorescence in situ hybridization (FISH) and for relevant protein expression by immunohistochemical analysis, also taking advantage of phosphorylated (p-) antibodies. Moreover, levels of ALK and MET mRNA were also determined by real-time polymerase chain reaction and Western blot analysis for downstream activation pathways involving p-MET, p–protein kinase B, p–mitogen-activated protein kinase, p-SRC proto-oncogene tyrosine-protein kinase, and p–focal adhesion kinase (p-FAK). Results MET amplification was detected by FISH in 25 of 98 PSCs (25.6%) and ALK amplification (but not the relevant rearrangement) was found in 16 of 98 (16.3%), with all ALK -amplified tumors also showing MET amplification ( p MET amplification without any ALK gene alteration. ALK protein expression was always lacking, whereas MET and p-MET were confined to the relevant amplified tumors only. Increased levels of ALK and MET mRNA were detectable in tumors with no direct relationship between mRNA content, protein expression, or alterations detected by FISH. Western blot assays showed complete activation of downstream signal pathways up to p-SRC proto-oncogene tyrosine-protein kinase, and p–focal adhesion kinase recruitment in MET and ALK– coamplified tumors only, whereas isolated MET amplification, MET and ALK borderline amplification (5% – 10% of tumor cells with ≥15 copies of the relevant gene), or negative tumors showing eusomy or chromosome polysomy were confined to p–mitogen-activated protein kinase, p–protein kinase B, and/or p-MET activation. Multivariate survival analysis pushed a higher percentage of MET altered cells or a higher value of MET copy gain per cell to marginally emerge for overall survival ( p = 0.140) and disease-free survival ( p = 0.060), respectively. Conclusions ALK and MET seemed to act as synergistic, nonrandom coactivators of downstream signal when coamplified in a subset of patients with PSC, thus likely suggesting a combined mechanism of oncogene addiction. These alterations could be a suitable target for therapy based on specific inhibitors.

The efficacy of computed tomography (CT) screening for early lung cancer detection in heavy smokers is currently being tested by a number of randomized trials. Critical issues remain the frequency of unnecessary treatments and impact on mortality, indicating the need for biomarkers of aggressive disease. We explored microRNA (miRNA) expression profiles of lung tumors, normal lung tissues and plasma samples from cases with variable prognosis identified in a completed spiral-CT screening trial with extensive follow-up. miRNA expression patterns significantly distinguished: ( i ) tumors from normal lung tissues, ( ii ) tumor histology and growth rate, ( iii ) clinical outcome, and ( iv ) year of lung cancer CT detection. Interestingly, miRNA profiles in normal lung tissues also displayed remarkable associations with clinical features, suggesting the influence of a permissive microenvironment for tumor development. miRNA expression analyses in plasma samples collected 1–2 y before the onset of disease, at the time of CT detection and in disease-free smokers enrolled in the screening trial, resulted in the generation of miRNA signatures with strong predictive, diagnostic, and prognostic potential (area under the ROC curve ≥ 0.85). These signatures were validated in an independent cohort from a second randomized spiral-CT trial. These results indicate a role for miRNAs in lung tissues and plasma as molecular predictors of lung cancer development and aggressiveness and have theoretical and clinical implication for lung cancer management.

INTRODUCTION miRNAs play a role in the complex network of signaling between cancer cells and tumor microenvironment. We previously reported the identification of diagnostic miRNA signatures (MSC) based on 24-miRNAs in plasma samples of lung cancer patients detected by low dose computed tomography (LDCT) screening. MATERIAL and METHODS To evaluate the potential origin of the miRNAs of the diagnostic signature, we analyzed their expression by real-time or digital PCR in both cells and conditioned medium (CM) from different cell types of the lung microenvironment as well as in plasma samples of heavy smokers and patients. Lung tissues and cell-blocks were analyzed by miRNAs in situ hybridization. Modulation of miRNAs after in vitro treatments, known to induce changes associated with cancer progression, in different cell types was assessed and correlated to changes observed in circulating miRNAs signatures. RESULTS and DISCUSSION The analysis of 100 pre and post surgery plasma samples from 31 patients suggested a tumor-related origin of those miRNAs which significantly declined to basal levels after curative tumor resection. Other miRNAs (miR-126, miR-92a, miR-320, miR-28-3p, miR-486, miR-451, miR-16) remained deregulated afer surgery likely indicating a “host-related” origin and the persistence of a risk profile. Specific expression of mir-145 in fibroblasts, mir-126 in endothelial cells, mir-133 in skeletal muscle cells, mir-451 and 142-3p in hematopoietic cells was observed with a good degree of correlation between cellular and secreted miRNAs levels in each cell types (Pearson correlation range: 0.59-0.81). In vitro experiments comparing activated vs. resting neutrophils, showed that 17 out of the 24 miRNAs were concordantly modulated as in lung cancer patients' plasma (Pearson r=0.60, p CONCLUSION These findings suggest that circulating miRNAs are non cell-autonomous, may act in paracrine signalling and have a causative role in lung carcinogenesis and immunesuppression. Citation Format: Orazio Fortunato, Cristina Borzi, Massimo Milione, Giovanni Centonze, Davide Conte, Mattia Boeri, Massimo Moro, Federica Facchinetti, Luca Roz, Veronica Huber, Chiara Camisaschi, Chiara Castelli, Licia Rivoltini, Valeria Cancila, Claudio Tripodo, Ugo Pastorino, Gabriella Sozzi. Circulating mir-320 promotes immunosuppressive macrophages M2 phenotype associated with lung cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4981.

A fusion gene, echinoderm microtubule associated protein like 4 – anaplastic lymphoma kinase (EML4-ALK), with transforming activity has recently been identified in a subset of non-small cell lung cancer (NSCLC), but its pathogenetic, diagnostic, and therapeutic roles remain unclear. Both frequency and type of EML4-ALK transcripts were investigated by reverse transcription PCR in 120 frozen NSCLC specimens from Italy and Spain; non-neoplastic lung tissues taken far from the tumor were used as controls. In cases carrying the fusion transcript, we determined EML4-ALK gene and protein levels using fluorescence in situ hybridization, Western blotting, and immunoprecipitation. We also analyzed ALK protein levels in paraffin samples from 662 NSCLC specimens, including the 120 cases investigated in the molecular studies. EML4-ALK transcripts (variants 1 and 3) were detected in 9 of 120 NSCLC samples but were not specific for NSCLC since they were also found in non-cancerous lung tissues taken far from the tumor. Notably, no transcripts were detected in matching tumor samples from these patients. Fluorescence in situ hybridization analysis of cases expressing EML4-ALK transcripts showed that only a minority of cells harbored the EML4-ALK gene. None of these cases was found to express the EML4-ALK protein as examined by immunohistochemistry, Western blotting, and immunoprecipitation. The EML4-ALK transcript cannot be regarded as a specific diagnostic tool for NSCLC. Our results show therefore that the causal role and value of EML4-ALK as a therapeutic target remain to be defined.

Background Research efforts for the management of cancer, in particular for lung cancer, are directed to identify new strategies for its early detection. MicroRNAs (miRNAs) are a new promising class of circulating biomarkers for cancer detection, but lack of consensus on data normalization methods has affected the diagnostic potential of circulating miRNAs. There is a growing interest in techniques that allow an absolute quantification of miRNAs which could be useful for early diagnosis. Recently, digital PCR, mainly based on droplets generation, emerged as an affordable technology for precise and absolute quantification of nucleic acids. Results In this work, we described a new interesting approach for profiling circulating miRNAs in plasma samples using a chip-based platform, the QuantStudio 3D digital PCR. The proposed method was validated using synthethic oligonucleotide at serial dilutions in plasma samples of lung cancer patients and in lung tissues and cell lines. Conclusion Given its reproducibility and reliability, our approach could be potentially applied for the identification and quantification of miRNAs in other biological samples such as circulating exosomes or protein complexes. As chip-digital PCR becomes more established, it would be a robust tool for quantitative assessment of miRNA copy number for diagnosis of lung cancer and other diseases. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-2097-9) contains supplementary material, which is available to authorized users.

We evaluated the aberrant promoter methylation profile of a panel of 3 genes in DNA from tumor and sputum samples, in view of a complementary approach to spiral computed tomography (CT) for early diagnosis of lung cancer. The aberrant promoter methylation of RARbeta2, p16(INK4A) and RASSF1A genes was evaluated by methylation-specific PCR in tumor samples of 29 CT-detected lung cancer patients, of which 18 had tumor-sputum pairs available for the analysis, and in the sputum samples from 112 cancer-free heavy smokers enrolled in a spiral CT trial. In tumor samples from 29 spiral CT-detected patients, promoter hypermethylation was identified in 19/29 (65.5%) cases for RARbeta2, 12/29 (41.4%) for p16(INK4A) and 15/29 (51.7%) for RASSF1A. Twenty-three of twenty-nine (79.3%) samples of the tumors exhibited methylation in at least 1 gene. In the sputum samples of 18 patients, methylation was detected in 8/18 (44.4%) for RARbeta2 and 1/18 (5%) for both RASSF1A and p16(INK4A). At least 1 gene was methylated in 9/18 (50%) sputum samples. Promoter hypermethylation in sputum from 112 cancer-free smokers was observed in 58/112 (51.7%) for RARbeta2 and 20/112 (17.8%) for p16, whereas methylation of the RASSF1A gene was found in only 1/112 (0.9%) sputum sample. Our study indicates that a high frequency of hypermethylation for RARbeta2, p16(INK4A) and RASSF1A promoters is present in spiral CT-detected tumors, whereas promoter hypermethylation of this panel of genes in uninduced sputum has a limited diagnostic value in early lung cancer detection.

Recent studies have demonstrated the possibility to detect genetic changes in plasma DNA of cancer patients. The goal of this study was to validate a panel of molecular markers for lung cancer detection in plasma DNA. Three markers, p53, FHIT and microsatellite alterations at loci on chromosome 3, were used to detect mutations in tumor and plasma DNA of 64 stage I-III non small cell lung cancer patients. p53 mutations were studied by direct sequencing of exons 5 through 8 in tumor DNA and by plaque hybridization assay and sequencing in plasma DNA. Allelic losses were evaluated by fluorescent PCR in tumor and plasma DNA. p53 genomic mutations were detected in 26 (40.6%) of 64 tumor DNA samples and the identical mutation was identified in plasma of 19 (73.1%) of them. Microsatellite alterations at FHIT and 3p loci were observed in 40 (62.5%) tumors and in 23 (35.9%) plasma samples. Of the 40 patients showing microsatellite alterations in tumors, 19 (47.5%) displayed the same change in plasma DNA. At least 1 of the 3 genetic markers (p53, FHIT and 3p) was altered in plasma of 51.6% of all patients and 60.7% of stage I patients. Moreover, genetic markers in plasma identified 29 of 45 (64.4%) of all stages and 15 of 22 (68.2%) of stage I patients whose tumors had an alteration. These results provide the proof of principle that plasma DNA alterations are tumor-specific in most cases and support blood testing as a noninvasive strategy for early detection.

Analysis of molecular markers in biological fluids has been proposed as a tool for early detection and monitoring of cancer. Circulating plasma DNA concentrations have been found to be higher in cancer patients than in cancer-free control subjects, but little is known about the effect of specimen storage on plasma DNA concentrations. Here we investigated the impact of long-term storage of both plasma samples and purified plasma DNA on the reproducibility of plasma DNA quantification as determined using real-time polymerase chain reaction analysis. The analysis was performed on samples from a subset of 34 lung cancer patients and 28 matched control subjects selected from 200 subjects in our previously published case-control study and from 117 cancer-free smokers enrolled in a lung cancer screening program. Two samples of plasma and isolated DNA were assessed for each patient, with a median of 41 months between the first and second assessments for participants in the case-control study and 9 months for participants in the screening study. DNA levels declined substantially between the two assessments at an average rate of approximately 30% per year. These data provide valuable information for the rational planning of retrospective studies of banked series of biological samples, particularly if collected over a long period of time, as can occur in large clinical trials.

Introduction FHIT and p53 are the two most commonly altered tumor suppressor genes in lung cancer, and their molecular status regulates sensitivity to anticancer drugs. Although their functions are independent, there is evidence that their pathways might be interconnected, but little is known at the molecular level. Methods Microarray profiling of FHIT-transduced lung cancer cells and modulation of FHIT levels by RNA interference in human bronchial cells were used to generate a signature of FHIT-regulated transcripts. Expression of these genes was evaluated by real-time polymerase chain reaction in 55 primary lung cancer samples characterized for FHIT and p53 expression by immunehistochemistry. Results A signature of FHIT-transcripts, particularly enriched in genes involved in cell cycle control, was identified. This signature showed overlap with p53-regulated genes, indicating possible crosstalk between these proteins. Consistently, transcriptional deregulation after FHIT modulation was higher in p53-negative cells. In primary lung cancers, inactivation of either gene was detected in 48 of 55 cases (87%) and both genes in 23 of 55 (42%) cases, confirming the central role of these pathways. Primary tumors with inactivation of both FHIT and p53 displayed the strongest deregulation of growth-related pathways with high levels of expression of CCNB1, BUB1, CDC6, TOP2A, MCM6, and CENPF. Conclusions FHIT and p53 seem to rely on common mediators, and inactivation of both genes results in prominent deregulation of growth-related pathways in lung cancer cell lines and primary tumors. This reveals crosstalk between these proteins and suggests a possible distinctive phenotype for tumors with inactivation of both genes.

In today’s modern societies the lack of computers feared by Asimov is not an imminent danger thanks to the advance of mobile technology in the last two decades. According to several market surveys, sales of handheld devices, especially smartphones, are growing at an incredibly fast rate and are expected to exceed those of any other electronic device by the end of 2011 1. This success is far from surprising, as today’s handheld devices feature high computational power and provide a wide range of applications that go beyond the traditional use of a phone. Examples are mHealth, a term coined by Istepanian et al. (2005) that refers to the use of mobile applications in healthcare, and augmented reality, defined by Azuma (1997) as a variation of virtual reality that “allows the user to see the real world, with virtual objects superimposed upon or composited with the real world”. Essentially, handheld devices fit in a pocket and provide most of the functionalities of a bulky computer. Their small size, however, is a mixed blessing, as it imposes serious limitations on usability, which is the focus of this chapter. In particular, we discuss two important aspects of the interaction with handheld devices, namely context-awareness and text entry.

The present study is aimed to design a prototype of hybrid silicon-muscle cell junction, analog to an artificial neuromuscular junction prototype and relevant to the development of advanced neuro-prostheses and bionic systems. The device achieves focal Electric Capacitive Stimulation (ECS) by coupling of single cells and semiconductors, without electrochemical reaction with the substrate. A voltage change applied to a stimulation spot beneath an electrogenic cell leads to a capacitive current (charge accumulation) that opens voltage-gated ion channels in the membrane and generates an action potential. The myo-electronic junction was employed to chronically stimulate muscle cells via ECS and to induce cytosolic calcium transients in myotubes, fibers isolated from mouse FDB (fast [Ca 2+ ] i transients) and surprisingly also in undifferentiated myoblasts (slow [Ca 2+ ] i waves). The hybrid junction elicited, via chronic ECS, a differential reprogramming of single muscle cells by inducing early muscle contraction maturation and plasticity effects, such as NFAT-C3 nuclear translocation. In addition, in the presence of agrin, chronic ECS induced a modulation of AchR clustering which simulates in vitro synaptogenesis. This methodology can coordinate the myogenic differentiation, thus offering direct but non-invasive single cell/wiring, providing a platform for regenerative medicine strategies.

Rationale: Fragile histidine triad (FHIT) is a tumor suppressor gene involved in the pathogenesis of lung cancer. Objectives: The purpose of this study was to investigate the different molecular alterations leading to the inactivation of FHIT gene function and to validate their use as biomarkers of risk for progression of the disease in patients belonging to the multicentric European study for the Early detection of Lung Cancer (EUELC) who were resected for early-stage lung tumors. Methods: FHIT immunostaining was performed on 305 tumor samples. Themethylation status of FHIT promoterwas assessed by nested methylation-specific polymerase chain reaction (MSP-PCR) in 232 tumor and 225 normal lung samples ofwhich a subset of 187 patients had available normal/tumorDNA pairs. Loss of heterozygosity (LOH) at the FHIT locus was analyzed in 202 informative cases by D3S1300 and D3S1234 microsatellite markers. Measurements and Main Results: Lost or reduced FHIT expression was found in 36.7 and 75.7% of the tumor samples, respectively. Methylation of the FHIT promoter was found in 36.7%of tumor and 32.7% of normal lung samples, whereas LOH was detected in 61.9% of the tumors. A strong association with complete loss of FHIT expression was presentwhenmethylation and LOHwere analyzed together (P5 0.0064). Loss of FHIT protein expression was significantly more frequent in squamous cell carcinoma histotype (P , 0.0001) and in smokers (P5 0.008). FHIT methylation in normal lung was associated with an increased risk of progressive disease (OR, 2.27; P 5 0.0415). Conclusions:Our results indicate thatdifferentmolecularmechanisms interplay to inactivate FHIT expression and support the proposition that FHIT methylation in normal lung tissue could represent a prognostic marker for progressive disease.

Rearrangements involving the ALK gene define two distinct entities in the new 2008 WHO classification of lymphoid neoplasms, i.e. ALK+ anaplastic large cell lymphoma and a rare subset of ALK+ diffuse large B-cell lymphoma. Recently, rearrangements involving ALK and the echinoderm microtubule associated protein-like 4 (EML4) gene were described as a specific genetic alteration in about 6% of non-small cell lung cancer (NSCLC). We investigated the expression of EML4-ALK mRNA and protein in 51 reactive and 58 neoplastic lymphoid tissues. EML4-ALK transcripts were detected in 3/51 (5.9%) of reactive lymphoid tissues and 12/58 (20.7%) of lymphomas of different categories, including follicular lymphoma, diffuse large B-cell lymphoma and Hodgkin’s disease. Notably, none of these cases expressed the EML4-ALK fusion protein at Western blotting samples and immunohistochemistry. These results indicate that EML4-ALK rearrangements are not specific of NSCLC and raise yet unsolved questions about their role in promoting human neoplasms.

Free circulating plasma DNA has emerged as a potential biomarker for early lung cancer detection. In a previous case-control study we have shown that high levels of plasma DNA are a strong risk factor for lung cancer.To assess the diagnostic performance and prognostic value of plasma DNA levels in a cohort of 1,035 heavy smokers monitored by annual spiral computed tomography (CT) for 5 years.Plasma DNA levels were determined through real-time quantitative PCR at baseline and at time of lung cancer diagnosis. Screening performance of the assay was calculated through the area under the receiver-operating characteristic curve (AUC-ROC). Kaplan-Meier analyses were computed for association with prognosis.Median baseline concentration of plasma DNA was not different in individuals who developed CT-detected lung cancers in the 5-year period (n = 38) versus cancer-free control subjects (AUC-ROC, 0.496; P = 0.9330), and only slightly higher at the time of cancer diagnosis (AUC-ROC, 0.607; P = 0.0369). At surgery, plasma DNA was higher in tumors detected at baseline (AUC-ROC, 0.80; P0.0001) and in Stage II to IV tumors detected during the first 2 years of screening (AUC-ROC, 0.87; P0.0001). A longitudinal study of plasma DNA levels showed increased values approaching to lung cancer diagnosis (P = 0.0010). Higher plasma DNA was significantly associated with poorer 5-year survival (P = 0.0066).Baseline assessment of plasma DNA level does not improve the accuracy of lung cancer screening by spiral CT in heavy smokers. Higher levels of plasma DNA at surgery might represent a risk factor for aggressive disease.

In this paper we report a set of results extracted from our experience in using PDAs and 3G multimedia cellular phones in Cultural Heritage (CH) through projects covering an activity spanning 10 years. In particular, we focus on a interesting variant of context-awareness, namely attention-awareness, derived from our more recent ICT European project, named Agamemnon. Such concepts are essential to the development of next generation mobile applications in cultural heritage and in other comparable fields. We present an experiment tackling this new issue by exploiting image recognition technology and we deeply analyze advantages and drawbacks of this approach, showing some preliminary results.

s / Gait & Posture 42S (2015) S1–S27 S15 Discussion: The common longer dual support time strategy in PwMS, which is independent of walking speed [3], results in an inappropriate activation of themuscle synergies that affects PwMS gait. In fact, despite anticipated M1 activation, PwMS showed a late dorsiflexion peak at contro-lateral heel strike that could be the consequence of reduced activation in force and time of the plantarflexor module (M1) in mid and terminal stance (Fig. 1) that likely compromises the proper control of tibial advancement. This may result in a non functional ankle position at the beginning of propulsive phase and consequently in a push-off deficit (Fig. 1). These results will allow a tailored rehabilitation and training in PwMS.