Efficacy and Safety of Alprostadil in Patients with Peripheral Arterial Occlusive Disease Fontaine Stage IV: Results of a Placebo Controlled Randomised Multicentre Trial (ESPECIAL).

OBJECTIVES: The aim was to assess the efficacy and safety of alprostadil in patients with peripheral arterial occlusive disease (PAOD) Fontaine Stage IV. METHODS: This was a multinational, prospective, randomised, double blind, placebo controlled, parallel group trial. Patients with Stage IV PAOD were equally randomised to either 4 weeks of alprostadil treatment twice daily or to placebo treatment twice daily. The primary efficacy variables were the rate of complete healing of all necrosis and ulceration 12 weeks after the end of treatment and the frequency of major amputations 24 weeks after the end of treatment. RESULTS: A total of 840 patients were randomised between 2004 and 2013. At baseline, no major differences between treatment groups were found. The rate of "complete healing" was 18.4% in patients on alprostadil and 17.2% in patients on placebo. The rates of "major amputations" were 12.6% in patients on alprostadil and 14.6% in patients on placebo. The adjusted difference between alprostadil and placebo including their 95% confidence intervals was 1.1 (-4.0 to 6.3) for "complete healing" and -2.1 (-6.7 to 2.5) for "major amputations." In the subgroup of diabetic patients the rates of major amputations were numerically lower in the alprostadil than placebo group (10.6% vs. 17.4%). Within the total cohort a non-significant difference in "decrease in ulcer area ≥50%" was reached in 30.2% of patients on alprostadil and in 24.3% of patients on placebo at end of treatment. CONCLUSIONS: In this study, superiority of alprostadil over placebo could not be shown. Nevertheless, a slight numerical but not clinically relevant advantage for alprostadil emerged from the "area decrease of ulcers by ≥ 50%," indicating that a healing effect may have started. The results have to be considered in the light of several limitations in study design and conduct.

ITC recommendations for transporter kinetic parameter estimation and translational modeling of transport-mediated PK and DDIs in humans.

This white paper provides a critical analysis of methods for estimating transporter kinetics and recommendations on proper parameter calculation in various experimental systems. Rational interpretation of transporter-knockout animal findings and application of static and dynamic physiologically based modeling approaches for prediction of human transporter-mediated pharmacokinetics and drug-drug interactions (DDIs) are presented. The objective is to provide appropriate guidance for the use of in vitro, in vivo, and modeling tools in translational transporter science.

Detection of epidermal growth factor receptor gene mutations in cytology specimens from patients with non-small cell lung cancer utilising high-resolution melting amplicon analysis.

BACKGROUND: Activating epidermal growth factor receptor (EGFR) mutations have been implicated in non-small cell lung cancer (NSCLC), and have also been clinically correlated with patient sensitivity to targeted EGFR inhibitors. AIM: To describe a technique for determining EGFR mutation status on archival fine needle aspirate (FNA) specimens from advanced NSCLC patients. METHODS: Eleven archival FNA slides from patients with advanced NSCLC were examined for diagnostic material to identify tumour cell-enriched regions. EGFR mutation status was determined using a slide-scrape DNA extraction protocol of selected tumour cell regions on the smear slides, followed by real time PCR and high resolution melt analysis (HRMAA) of EGFR exons 18, 19, 20, and 21, followed by sequence analysis. RESULTS: All DNA samples were successfully amplified by PCR. Three adenocarcinoma patient samples contained EGFR mutations in exon 19 (L747-P753insS). One of the three had an additional exon 19 mutation (A755D). CONCLUSIONS: Archival cytology slides from patients with NSCLC can be used to determine EGFR mutation status by PCR, HRMAA, and sequencing. The ability to use archival cytology slides greatly increases the potential material available for molecular analysis in diagnosis and selection of patients for targeted therapeutic agents.

Immediate on-site interpretation of fine-needle aspiration smears: a cost and compensation analysis.

BACKGROUND: A significant body of literature exists supporting the cost effectiveness of fine-needle aspiration (FNA) cytology in the work-up of patients with potential neoplastic disease. Several authorities have stated that immediate, on-site smear evaluation by cytopathologists optimizes diagnostic accuracy and minimizes the technique's insufficiency rate. This favorable effect on FNA diagnostic accuracy is most pronounced for deep body sites, where FNA is guided by computed tomography (CT), ultrasound, bronchoscopy, or endoscopy. Little data exist regarding whether compensation from Medicare is adequate to support the pathologist in this endeavor compared with other potentially more remunerative activities, including routine surgical pathology sign-out, nongynecologic cytopathology sign-out, and frozen section consultation. METHODS: The authors studied a series of 142 fine-needle aspirates with immediate, on-site evaluations performed under a variety of clinical settings. These included bronchoscopic, endoscopic, ultrasound-guided, and CT-guided biopsies along with palpation-directed biopsies performed by either cytopathologists or clinicians. For these aspirates, total pathologist attendance time was calculated and correlated with guidance technique, target organ, location where aspirate was performed, and nature of aspirator. Fifty frozen section evaluations were timed similarly. For comparison purposes, cytopathologists' costs were calculated using the 80th percentile pay level of an associate professor with full-time clinical duties. Medicare rate schedules were used to calculate compensation. Including salary and benefits, the pathologist cost was approximately $88.83 per hour. RESULTS: On average, an intraprocedural FNA evaluation for a CT-guided biopsy required 48.7 minutes, an ultrasound-guided biopsy required 44.4 minutes of pathologist time, an endoscopic procedure required 56.2 minutes, a bronchoscopic procedure required 55.3 minutes, a clinic aspirate performed by a pathologist required 42.5 minutes, and a clinic FNA performed by a clinician required 34.7 minutes. The average frozen section required 15.7 minutes of pathologist time for performance and interpretation. With the exception of FNA performed in clinic by the cytopathologist, time costs exceeded compensation by $40-50 per procedure. Clinic aspirates performed by a clinician and immediately evaluated by a pathologist resulted in a deficit of approximately $18 over actual time cost. CONCLUSIONS: From the current data, it appears that intraprocedural consultations by cytopathologists for CT-guided, ultrasound-guided, bronchoscopic, or endoscopic procedures are compensated insufficiently by current Medicare compensation schedules using the CPT code 88172 for on-site evaluation. Only when the cytopathologist personally performs the aspirate and immediately interprets it (CPT codes 88172 and 88170) does the Medicare payment adequately compensate for professional services.

Comprehensive kinetic analysis of influenza hemagglutinin-mediated membrane fusion: role of sialate binding.

The data of Danieli et al. (J. Cell Biol. 133:559-569, 1996) and Blumenthal et al. (J. Cell Biol. 135:63-71, 1996) for fusion between hemagglutinin (HA)-expressing cells and fluorescently labeled erythrocytes has been analyzed using a recently published comprehensive mass action kinetic model for HA-mediated fusion. This model includes the measurable steps in the fusion process, i.e., first pore formation, lipid mixing, and content mixing of aqueous fluorescent markers. It contains two core parameters of the fusion site architecture. The first is the minimum number of aggregated HAs needed to sustain subsequent fusion intermediates. The second is the minimal number of those HAs within the fusogenic aggregate that must undergo a slow "essential" conformational change needed to initiate bilayer destabilization. Because the kinetic model has several parameters, each data set was exhaustively fitted to obtain all best fits. Although each of the data sets required particular parameter ranges for best fits, a consensus subset of these parameter ranges could fit all of the data. Thus, this comprehensive model subsumes the available mass action kinetic data for the fusion of HA-expressing cells with erythrocytes, despite the differences in assays and experimental design, which necessitated transforming fluorescence dequenching intensities to equivalent cumulative waiting time distributions. We find that HAs bound to sialates on glycophorin can participate in fusion as members of the fusogenic aggregate, but they cannot undergo the essential conformational change that initiates bilayer destabilization, thus solving a long-standing debate. Also, the similarity in rate constants for lipid mixing and content mixing found here for HA-mediated fusion and by Lee and Lentz (Proc. Natl. Acad. Sci. U.S.A. 95:9274-9279, 1998) for PEG-induced fusion of phosphatidylcholine liposomes supports the idea that subsequent to stable fusion pore formation, the evolution of fusion intermediates is determined more by the lipids than by the proteins.

Cell block preparation as an adjunctive diagnostic technique in ThinPrep monolayer preparations: a case report.

The cell block preparation, which increases cellular yield by capturing any small tissue fragments in a fluid specimen, has been used to define the histologic pattern of the cells in question. This report describes a case in which processing a cell block on residual ThinPrep Pap Test material, using a thrombin-based technique, was useful in augmenting the diagnosis of a glandular lesion.

Deployment of membrane fusion protein domains during fusion.

It is clear that both viral and intracellular membrane fusion proteins contain a minimal set of domains which must be deployed at the appropriate time during the fusion process. An account of these domains and their functions is given here for the four best-described fusion systems: influenza HA, sendai virus F1, HIV gp120/41 and the neuronal SNARE core composed of synaptobrevin (syn), syntaxin (stx) and the N- and C-termini of SNAP25 (sn25), together with the Ca(2+)binding protein synaptotagmin (syt). Membrane fusion begins with the binding of the virion or vesicle to the target membrane via receptors. The committed step in influenza HA- mediated fusion begins with an aggregate of HAs (at least eight) with some of their HA2 N-termini, a.k.a. fusion peptides, embedded into the viral bilayer (Bentz, 2000 a). The hypothesis presented in Bentz (2000 b) is that the conformational change of HA to the extended coiled coil extracts the fusion peptides from the viral bilayer. When this extraction occurs from the center of the site of restricted lipid flow, it exposes acyl chains and parts of the HA transmembrane domains to the aqueous media, i.e. a hydrophobic defect is formed. This is the 'transition state' of the committed step of fusion. It is stabilized by a 'dam' of HAs, which are inhibited from diffusing away by the rest of the HAs in the aggregate and because that would initially expose more acyl chains to water. Recruitment of lipids from the apposed target membrane can heal this hydrophobic defect, initiating lipid mixing and fusion. The HA transmembrane domains are required to be part of the hydrophobic defect, because the HA aggregate must be closely packed enough to restrict lipid flow. This hypothesis provides a simple and direct coupling between the energy released by the formation of the coiled coil to the energy needed to create and stabilize the high energy intermediates of fusion. Several of these essential domains have been described for the viral fusion proteins SV5 F1 and HIV gp120/41, and for the intracellular SNARE fusion system. By comparing these domains, we have constructed a minimal set which appears to be adequate to explain how the conformational changes can produce a successful fusion event, i.e. communication of aqueous compartments.

New technologies in gynecologic cytology.

New technologies are trying to make significant changes in the way we perform the Pap smear test, a test that has undergone no significant modification since its inception. Efforts have focused on improving sampling with liquid-based methods, improving detection of abnormal cells with automated screening and improving the detection of human papillomavirus with assays practical for use in routine clinical material. This article reviews the performance of the new technologies to date, with an emphasis on those that are currently approved for cervical cytology testing. In the aggregate, these technologies can have a positive impact on cervical disease detection and intervention.

Quantification of HER2 oncoprotein in fine-needle aspirates of the breast.

Measurements of either HER2 gene overexpression or its gene-coded protein (p185) are clinically useful for predicting prognosis in breast cancer. The measurements are also useful for identifying metastatic breast cancer patients who may benefit from Herceptin treatment. Since fine needle aspiration (FNA) of the breast has become an increasingly popular technique for obtaining tissue specimens, we have developed a sensitive method to quantify p185 in the aspirate. For this procedure, p185 from the cell pellet of FNA is extracted with a buffer containing Triton X-100, and the p185 is measured with an enzyme immunoassay. Most of the malignant breast tumors (N=7) in this study were associated with elevated p185 concentrations (6/7, 319+/-222 U/mg), compared to the p185 concentrations in normal breast tissue (42.8+/-35 U/mg, N=47) or benign lesions (43.1+/-20.2 U/mg, N=22). Quantification of p185 in FNA may improve the assessment of breast cancer patients, revealing whether they are at high risk and may benefit from Herceptin treatment.

Membrane fusion mediated by coiled coils: a hypothesis.

A molecular model of the low-pH-induced membrane fusion by influenza hemagglutinin (HA) is proposed based upon the hypothesis that the conformational change to the extended coiled coil creates a high-energy hydrophobic membrane defect in the viral envelope or HA expressing cell. It is known that 1) an aggregate of at least eight HAs is required at the fusion site, yet only two or three of these HAs need to undergo the "essential" conformational change for the first fusion pore to form (Bentz, J. 2000. Biophys. J. 78:000-000); 2) the formation of the first fusion pore signifies a stage of restricted lipid flow into the nascent fusion site; and 3) some HAs can partially insert their fusion peptides into their own viral envelopes at low pH. This suggests that the committed step for HA-mediated fusion begins with a tightly packed aggregate of HAs whose fusion peptides are inserted into their own viral envelope, which causes restricted lateral lipid flow within the HA aggregate. The transition of two or three HAs in the center of the aggregate to the extended coiled coil extracts the fusion peptide and creates a hydrophobic defect in the outer monolayer of the virion, which is stabilized by the closely packed HAs. These HAs are inhibited from diffusing away from the site to admit lateral lipid flow, in part because that would initially increase the surface area of hydrophobic exposure. The other obvious pathway to heal this hydrophobic defect, or some descendent, is recruitment of lipids from the outer monolayer of the apposed target membrane, i.e., fusion. Other viral fusion proteins and the SNARE fusion protein complex appear to fit within this hypothesis.

Minimal aggregate size and minimal fusion unit for the first fusion pore of influenza hemagglutinin-mediated membrane fusion.

The data of Melikyan et al. (J. Gen. Physiol. 106:783, 1995) for the time required for the first measurable step of fusion, the formation of the first flickering conductivity pore between influenza hemagglutinin (HA) expressing cells and planar bilayers, has been analyzed using a new mass action kinetic model. The analysis incorporates a rigorous distinction between the minimum number of HA trimers aggregated at the nascent fusion site (which is denoted the minimal aggregate size) and the number of those trimers that must to undergo a slow essential conformational change before the first fusion pore could form (which is denoted the minimal fusion unit). At least eight (and likely more) HA trimers aggregated at the nascent fusion site. Remarkably, of these eight (or more) HAs, only two or three must undergo the essential conformational change slowly before the first fusion pore can form. Whether the conformational change of these first two or three HAs are sufficient for the first fusion pore to form or whether the remaining HAs within the aggregate must rapidly transform in a cooperative manner cannot be determined kinetically. Remarkably, the fitted halftime for the essential HA conformational change is roughly 10(4) s, which is two orders of magnitude slower than the observed halftime for fusion. This is because the HAs refold with distributed kinetics and because the conductance assay monitored the very first aggregate to succeed in forming a first fusion pore from an ensemble of hundreds or thousands (depending upon the cell line) of fusogenic HA aggregates within the area of apposition between the cell and the planar bilayer. Furthermore, the average rate constant for this essential conformational change was at least 10(7) times slower than expected for a simple coiled coil conformational change, suggesting that there is either a high free energy barrier to fusion and/or very many nonfusogenic conformations in the refolding landscape. Current models for HA-mediated fusion are examined in light of these new constraints on the early structure and evolution of the nascent fusion site. None completely comply with the data.

Acid-fast-positive Legionella pneumophila: a possible pitfall in the cytologic diagnosis of mycobacterial infection in pulmonary specimens.

The acid-fast stain is commonly used in the rapid cytologic assessment of bronchoalveolar lavage (BAL) fluid to detect pulmonary mycobacterial infections, particularly in immunocompromised patients. The identification of acid-fast, rod-shaped organisms may be taken as presumptive evidence of such an infection, in the appropriate clinical setting. However, this determination is made less specific by the occasional acid-fast positivity of microorganisms other than mycobacteria. We report on the occurrence of a fatal pneumonia caused by acid-fast positive Legionella pneumophila detected by BAL. This is a potential pitfall in the rapid diagnosis of pulmonary mycobacterial infections.

[Utilization of services at the Berlin Center of Ambulatory Rehabilitation]. / Inanspruchnahme von Leistungen im Berliner Zentrum für ambulante Rehabilitation.

The Center for Ambulatory Rehabilitation (ZaR) in Berlin provides rehabilitative services for orthopedic and neurological patient problems offering a rehabilitation program that is flexible, individually adapted and close to the patient's home. This paper analyzes the development of utilization of the ZaR using patient application, admission and discharge data for a one year period (April 1997 to March 1998). Treatment was started for 1,009 patients (mean age 51.1 years; 55% female). While mean duration of a treatment period was 28.5 days, overall utilization of the ZaR was 49%, being higher for the neurological department than for the orthopedic department (74% and 40%, respectively). The variety of patient problems treated was fairly small: more than two thirds of the cases treated were patients after stroke (ICD 430-438) in the neurological department and patients with back problems (ICD 721-724) in the orthopedic department, respectively. Acute care hospitals still play a minor role in referring patients to the ZaR. Referrals of many office-based physicians suggest that the ZaR will achieve its intention to provide rehabilitative services close to the patient's home.

Diagnostic accuracy in fine-needle aspiration of soft tissue and bone lesions. Influence of clinical history and experience.

Interpretation of soft tissue and bone lesions with fine-needle aspiration (FNA) is difficult. We determined whether clinical history or experience improves diagnostic accuracy of FNA interpretation of these lesions. Four cytopathologists with varying degrees of experience retrospectively reviewed 89 soft tissue and bone lesions, initially without knowledge of clinical history and subsequently with knowledge of clinical history. Each time, the pathologist rendered a precise diagnosis (histogenetic classification, e.g., angiosarcoma, lipoma) for each case and classified the lesion into one of 4 categories: benign, probably benign, probably malignant, or malignant. The proportion of correct precise diagnoses was calculated. Also, the numbers of correct and incorrect classifications of malignancy were used in standard relative operating characteristic (ROC) curve analysis. The area under the ROC curve was estimated to give an indicator of diagnostic accuracy. Knowledge of clinical history increased the proportion of correct diagnoses and the accuracy of the classification for all observers. Experienced observers more accurately classified lesions, and knowledge of clinical history particularly improved the diagnostic accuracy of less experienced observers.

DINAMO: interactive protein alignment and model building.

MOTIVATION: To facilitate the process of structure prediction by both comparative modeling and fold recognition, we describe DINAMO, an interactive protein alignment building and model evaluation tool that dynamically couples a multiple sequence alignment editor to a molecular graphics display. DINAMO allows the user to optimize the alignment and model to satisfy the known heuristics of protein structure by means of a set of analysis tools. The analysis tools return information to both the alignment editor and graphics model in the form of visual cues (color, shape), allowing for rapid evaluation. Several analysis tools may be employed, including residue conservation, residue properties (charge, hydrophobicity, volume), residue environmental preference, and secondary structure propensity. RESULTS: We demonstrate DINAMO by building a model for submission in the 3rd annual Critical Assessment of Techniques for Protein Structure Prediction (CASP3) contest. AVAILABILITY: DINAMO is freely available as a local application or Web-based Java applet at http://tito.ucsc.edu/dinamo

Improved quality-control detection of false-negative Pap smears using the Autopap 300 QC system.

Federally-mandated quality control (QC) in Papanicolaou (Pap) smear testing requires rescreening of 10% of negative smears, to include cases selected randomly as well as smears from patients that may have a higher risk for developing cervical cancer based on clinical information. FDA approval of NeoPath's AutoPap 300 QC system (NeoPath, Inc., Redmond, WA) allows practical QC rescreening of all negatives. We tested the ability of AutoPap to help increase identification of detection errors compared to random 10%/high-risk selection. From March 1-August 30, 1997, we utilized AutoPap/high-risk status to select cases for manual rescreen, and compared the rate of identification of primary screening errors to that for the preceding year using 10% random selection/high-risk status. Of 35,027 smears accessioned, 31,240 (89.1%) were screened as negative and 7,965 were selected for manual rescreen. Of these, 353 were determined to be abnormal. Most abnormals identified by this protocol were classified as atypical squamous or glandular cells of undetermined significance (ASCUS or AGUS). However, 59 low-grade squamous intraepithelial lesions (LSIL) and 13 high-grade squamous intraepithelial lesions (HSIL), many with few abnormal cells, were also identified. These results represented an increase in pickup rate of false negative due to detection errors of 2.3-, 2.8- and 5.6-fold for atypical squamous or glandular cells of undetermined significance, LSIL, and HSIL, respectively, when accounting for the volume differences over the time period measured. Our findings strongly support the conclusions drawn from clinical trials of the AutoPap that false negatives due to detection error can be significantly reduced when using AutoPap as part of a routine quality control program.

[Demographic and curative factors of need for rehabilitation--possibilities and limits of simple time series models]. / Demographische und kurative Bedingungsfaktoren des Reha-Bedarfs--Möglichkeiten und Grenzen von einfachen Zeitreihenmodellen.

Regarding rehabilitation demands, macro-analytical time-series models outline a method for the estimation of effective rehabilitation needs and for explaining the magnitude of the requirements. Their range is limited, in so far as they are unable to clarify the rehabilitation requirement regarding individual micro-level behavioural aspects. For the moment the rehabilitation requirements are hidden units in the models. Differing macro-dimensions have been gradually included in the analysis. The demographic parameters of the potential patients in need of rehabilitation are the fundamental starting point. Rehabilitation requirements are increasingly modelled by the magnitude of the curative requirements. These are characterised in the rehabilitation as "preliminary or follow up". Two examples of simple time-series models in rehabilitation--for the development of rehabilitation demand--illustrate empirically, which possibilities and boundaries are set in view of demographic and curative requirements by the interpretative range of the macro-concepts. What is methodically interesting with it, is how the analytical borders of such time-series models can experience a recognisable theoretical broadening, through a projection in real logistical facts--here in the interaction between prognosis and retrospection.

Pleomorphic lobular carcinoma of the breast: clinicopathologic features of 12 cases.

Pleomorphic lobular carcinoma (PLC) of the breast was recently identified as a histologic variant of infiltrating lobular carcinoma (ILC) with a poor prognosis. Twelve cases identified from a large series of breast carcinomas were studied retrospectively. Of 11 cases with adequate follow up, 9 were fatal. This was significantly worse than either infiltrating ductal carcinoma (IDC) or classical ILC (P < or = .002), even when stratified by axillary lymph node status. Among the fatal cases, the median survival time was 2.1 years, significantly shorter than that for classical lobular, but not ductal, carcinoma A distinctive pattern of in situ carcinoma, which has been described as PLC in situ, was identified in 7 of the 12 patients. This in situ component was composed of tumor cells with nuclear atypia, cytologically similar to the invasive tumor. Most PLCs lacked estrogen and progesterone receptors and stained with BRST-2, an antibody to gross cystic disease fluid protein-15, suggesting the presence of apocrine differentiation. In summary, PLC has many of the histologic features of ILC but has anaplastic nuclei, abundant cytoplasm, and apocrine differentiation. PLC is often aneuploid, usually lacks steroid receptors, and has a significantly poorer prognosis than does classical ILC.

DINAMO: a coupled sequence alignment editor/molecular graphics tool for interactive homology modeling of proteins.

Gaining functional information about a novel protein is a universal problem in biomedical research. With the explosive growth of the protein sequence and structural databases, it is becoming increasingly common for researchers to attempt to build a three-dimensional model of their protein of interest in order to gain information about its structure and interactions with other molecules. The two most reliable methods for predicting the structure of a protein are homology modeling, in which the novel sequence is modeled on the known three-dimensional structure of a related protein, and fold recognition (threading), where the sequence is scored against a library of fold models, and the highest scoring model is selected. The sequence alignment to a known structure can be ambiguous, and human intervention is often required to optimize the model. We describe an interactive model building and assessment tool in which a sequence alignment editor is dynamically coupled to a molecular graphics display. By means of a set of assessment tools, the user may optimize his or her alignment to satisfy the known heuristics of protein structure. Adjustments to the sequence alignment made by the user are reflected in the displayed model by color and other visual cues. For instance, residues are colored by hydrophobicity in both the three-dimensional model and in the sequence alignment. This aids the user in identifying undesirable buried polar residues. Several different evaluation metrics may be selected including residue conservation, residue properties, and visualization of predicted secondary structure. These characteristics may be mapped to the model both singly and in combination. DINAMO is a Java-based tool that may be run either over the web or installed locally. Its modular architecture also allows Java-literate users to add plug-ins of their own design.

Endoscopic ultrasound-guided real-time fine-needle aspiration: clinicopathologic features of 60 patients.

Diagnosis of lesions of the gastrointestinal tract and adjacent structures is possible using an imaging modality, endoscopic ultrasonography (EUS). Fine-needle aspiration (FNA) is a suitable and cost-effective technique for obtaining cytohistologic material to confirm the diagnosis. EUS is capable of both characterizing the lesion and then guiding the FNA under real-time (RT) ultrasound guidance using a through-the-scope needle aspiration system. The goal of this study was to determine the diagnostic accuracy of this technique and to describe the clinicopathologic features. Sixty patients underwent EUS-guided RTFNA of 64 lesions, including pancreas (n = 45), periluminal lymph nodes (n = 12), mediastinal and retroperitoneal masses (n = 4), and hepatobiliary masses (n = 3). Follow-up data were obtained by surgery, histopathology, or clinical course. Thirty-one lesions were malignant, eight were atypical/suspicious, 16 were non-neoplastic, and nine were non-diagnostic. Of the 55 lesions with sufficient material for interpretation, 54 had follow-up confirmation of the RTFNA diagnosis. The calculated sensitivity and specificity for malignancy was 90% and 100%, respectively. Diagnostic accuracy for malignancy was excellent for gastrointestinal associated lymph nodes (100%), mediastinal and retroperitoneal masses (100%), somewhat less so for pancreatic tumors (94%), and poor for hepatobiliary lesions (33%). EUS-guided RTFNA is accurate for sampling small gastrointestinal tract-associated lesions. EUS-guided RTFNA should be considered as a procedure of choice in selected patients when the results will influence management decisions.