Pediatric Specimens Research Articles

Respiratory virus identification by interval polymerase chain reaction testing in the southeastern United States

This study was designed to determine if testing the first ~40 nasal washings (interval) each month for 1 year, could be used as an epidemiologic tool for seasonality and prevalence of respiratory viruses such as human metapneumovirus in an adult and pediatric population in the southeastern United States. Results of interval polymerase chain reaction (PCR) testing of 469 specimens for 8 viruses were compared with our current procedures using PCR, culture, or respiratory synctial virus antigen for all 7435 specimens (routine). One hundred thirty-six viruses out of 469 specimens (29.0%) and 1,495 viruses out of 7,435 specimens (20.1%) were identified by interval and routine testing, respectively. Seasonal detection varied among viruses and to some degree between interval and routine testing. A higher percent of positives and dual infections were detected by interval testing of pediatric specimens, likely due to the use of PCR for viruses commonly seen in this population. Human metapneumovirus was detected in both pediatric and adult specimens between January and August. Interval testing can be used to provide a snapshot of prevalence and seasonality of respiratory viruses, although as currently designed they may not be sensitive enough to identify the beginning of a specific virus season. Exclusive use of interval PCR testing identified several dual infections, including human metapneumovirus, throughout most of the year in Florida. A rapid turnaround time to results translates into improved infection control and improved patient care.

Effect of patient Age on surgical outcomes for Graves’ disease: a case–control study of 100 consecutive patients at a high volume thyroid surgical center

ObjectiveTo compare outcomes between children (<18 yrs) and adults undergoing total thyroidectomy for Graves’ disease (GD) at a high volume, multidisciplinary thyroid center.Summary of background dataReported complication rates for children undergoing surgery for Graves’ disease are worse than for adults.Methods100 consecutive patients (32 children; 68 adults) who underwent total thyroidectomy for Graves’ disease (GD) by a high-volume endocrine surgery team from were compared.ResultsThe mean patient age was 9.7 yrs (range 3.4-17.9 yrs) in children versus 44.9 yrs (range 18.4-84.2 yrs) in adults. Operative times were longer in children (2.18 ± 0.08 hrs) than in adults (1.66 ± 0.03 hrs) (p = 0.003). Pediatric thyroid specimens averaged 38.6.0 ± 8.9 gm (range: 9–293 gm) and adult thyroid specimens averaged 48.0 ± 6.4 gm (range: 6.6-203 gm) (p = 0.34). Thyroid to body weight ratios were greater in children (0.94 ± 0.11 gm/kg) than adults (0.67 ± 0.8 gm/kg) (p = 0.05). In all patients, the hyperthyroid state resolved after surgery. There was no operative mortality, recurrence, or permanent hypoparathyroidism. Transient post-operative hypocalcemia requiring calcium infusion was greater in children than adults (6/32 vs. 1/68; p = 0.004). Transient recurrent laryngeal nerve dysfunction occurred in two children and in no adults (p = 0.32). Postoperative hematoma occurred in two adults and in no children (p = 0.46). The length of stay was longer for children (1.41 ± 0.12 days) than for adults (1.03 ±0.03 days) (p = 0.004).ConclusionSurgical management of GD is technically more challenging in children as evidenced by longer operative times. Whereas temporary hypocalcemia occurs more commonly in children than adults, the risks of major complications including disease recurrence, permanent hypoparathyroidism, recurrent laryngeal nerve injury, or neck hematoma were indistinguishable. These data suggest that excellent and equivalent outcomes can be achieved for GD surgery in children and adults when care is rendered by a high volume, endocrine surgery team.

Role of the faecolith in modern-day appendicitis

IntroductionThe prevailing view on appendicitis is that the main aetiology is obstruction owing to faecoliths in adults and lymphoid hyperplasia in children. Faecoliths on imaging studies are believed to correlate well with appendicitis.MethodsA retrospective chart review was conducted of 1,014 emergency appendicectomy patients between 2001 and 2011. Faecolith prevalence in adult and paediatric appendicectomy specimens with and without perforation was studied. The sensitivity and positive predictive value (PPV) of computed tomography (CT) for identifying faecoliths in the pathology specimen were examined.ResultsOverall, faecoliths were found in 18.1% (178/986) of appendicitis specimens and 28.6% (8/28) of negative appendicectomies. Faecolith prevalence for positive cases was 29.9% (79/264) in paediatric patients and 13.7% (99/722) in adults (p<0.05). Faecolith prevalence was 39.4% in perforated appendicitis but only 14.6% in non-perforated appendicitis (p<0.05). In adults, faecolith prevalence was 27.5% in perforated appendicitis and 12.0% in non-perforated appendicitis (p<0.05) while in paediatric patients, it was 56.1% in perforated appendicitis and 22.7% in non-perforated appendicitis (p=0.00). Sensitivity and PPV of preoperative CT in identifying faecoliths on pathology were 53.1% (86/162) and 44.8% (86/192) respectively.ConclusionsFaecolith prevalence is too low to consider the faecolith the most common cause of non-perforated appendicitis. Faecoliths are more prevalent in paediatric appendicitis than in adult appendicitis. Preoperative CT is an unreliable predictor of faecoliths in pathology specimens.

Characterizing biobank organizations in the U.S.: results from a national survey

BackgroundEffective translational biomedical research hinges on the operation of 'biobanks,' repositories that assemble, store, and manage collections of human specimens and related data. Some are established intentionally to address particular research needs; many, however, have arisen opportunistically, in a variety of settings and with a variety of expectations regarding their functions and longevity. Despite their rising prominence, little is known about how biobanks are organized and function beyond simple classification systems (government, academia, industry).MethodsIn 2012, we conducted the first national survey of biobanks in the U.S., collecting information on their origins, specimen collections, organizational structures, and market contexts and sustainability. From a list of 636 biobanks assembled through a multi-faceted search strategy, representatives from 456 U.S. biobanks were successfully recruited for a 30-minute online survey (72% response rate). Both closed and open-ended responses were analyzed using descriptive statistics.ResultsWhile nearly two-thirds of biobanks were established within the last decade, 17% have been in existence for over 20 years. Fifty-three percent listed research on a particular disease as the most important reason for establishment; 29% listed research generally. Other reasons included response to a grant or gift, and intent to centralize, integrate, or harmonize existing research structures. Biobank collections are extraordinarily diverse in number and types of specimens and in sources (often multiple) from which they are obtained, including from individuals, clinics or hospitals, public health programs, and research studies. Forty-four percent of biobanks store pediatric specimens, and 36% include postmortem specimens. Most biobanks are affiliated in one or multiple ways with other entities: 88% are part of at least one or more larger organizations (67% of these are academic, 23% hospitals, 13% research institutes). The majority of biobanks seem to fill a particular 'niche' within a larger organization or research area; a minority are concerned about competition for services, although many are worried about underutilization of specimens and long-term funding.ConclusionsEffective utilization of biobank collections and effective policies to govern their use will require understanding of the immense diversity found in organizational features, including the very different history and primary goals that many biobanks have.

Analytical measurement of serum 25-OH-vitamin D3, 25-OH-vitamin D2 and their C3-epimers by LC–MS/MS in infant and pediatric specimens

ObjectivesTo develop a simple and sensitive LC–MS/MS procedure for quantification of serum 25-OH-vitamin D3 (25-OH-D3), 25-OH-vitamin D2 (25-OH-D2), and their C3-epimers. MethodsSerum 25-OH-vitamin D metabolites were extracted with MTBE and quantified by LC–MS/MS. Commercially available calibrators and QC materials were employed. The ion-transition 401.2→365.2 was monitored for 25-OH-D3 and C3-epi-25-OH-D3, 407.2→371.3 for d6-25-OH-D3, 413.2→331.2 for 25-OH-D2 and C3-epi-25-OH-D2 and 419.2→337.1 for, d6-25-OH-D2. As a proof-of-principle, 25-OH-D3 and C3-epi-25-OH-D3 were quantified in 200 pediatric subjects (0–20years of age). Cholecalciferol supplements were examined as a potential source of C3-epimer. ResultsThe assay provided an LLOQ of ≤2.8nmol/L for all 25-OH-D metabolites, with a linear response up to 400nmol/L. The CV was <10% for 25-OH-D2/3 and <15% for C3-epi-25-OH-D3. C3-epi-25-OH-D3 was quantified in all subjects, with higher concentrations observed in infants ≤1year of age (11.44nmol/L vs. 4.4nmol/L; p<0.001). Within the first year of life, 25-OH-D3 concentrations increased linearly, while C3-epi-25-OH-D3 concentrations remained constant. At 12months of age, C3-epi-25-OH-D3 concentration dropped by almost 50% (11.4nmol/L in infants ≤1year of age vs. 5.4nmol/L in infants 1–2years of age; p<0.001). Liquid vitamin D3 supplements did not contain appreciable amounts of C3-epi-D3. ConclusionsThe proposed LC–MS/MS procedure is suitable for quantifying 25-OH-D3 metabolites. Although the C3-epimer is present in all pediatric subjects, it is significantly elevated in individuals ≤1year of age and drops at 12months of age. Oral vitamin D supplements are unlikely to be a significant source of C3-vitamin D epimer.

Otopathology in Osteogenesis Imperfecta

Osteogenesis Imperfecta (OI) is a genetic disorder of connective tissue matrix. OI is caused by mutations that affect type I collagen. The hearing loss in OI is characterized by onset in early adulthood and can be conductive, sensorineural, or mixed. To describe the temporal bone histopathology in 9 individuals with OI. Four adult, 1 pediatric, and 4 infant specimens were identified. Temporal bones were removed at autopsy and studied using light microscopy. All adults and 1 pediatric specimen showed otosclerotic lesions. The findings included examples of clinical, histologic, and cochlear otosclerosis. The temporal bones of infants showed delayed ossification of the endochondral layer of bone and of the ossicles. There were no infant specimens with otosclerotic lesions. Hearing loss in OI may be the result of clinical or cochlear otosclerosis. Fracture or atrophy of the ossicles may also be present in OI. A third unidentified mechanism of hearing loss may lead to cochlear degeneration. The described findings of otosclerotic lesions have implications for the observed heterogeneity of hearing loss patterns and for the surgical management of hearing loss in OI.

Whole slide images for primary diagnostics of paediatric pathology specimens: a feasibility study

IntroductionWhole slide images (WSI) have been used in many pathology applications such as teleconsultation, teaching and research, but not in primary diagnostics. The aim of this study was to test...

A New Algorithm and Panel Construction for Pediatric Leukemia Immunophenotyping Using 10-Color Flow Cytometry

Abstract 4799 Background:Rapid identification and quantification of abnormal cell populations in minimal specimen are crucial for diagnosis and longitudinal minimal residual disease (MRD) testing of childhood leukemia. So far, most standard immunophenotypic analyses are performed using antibody panels with up to five-colors and require high cell numbers. For infant and pediatric specimen, high-level multicolor analyses is highly desirable to gather sufficient data for initial diagnostic and follow up monitoring of pathologic populations. Objective:In this study, we aimed to establish a newly defined pediatric multicolor flow cytometric panel algorithm with high reliability yet minimal specimen requirement. Results:We defined a 10-color flow cytometric panel using the new violet laser dye “KromeOrange (KO)”. Applying CD45-KO/Side Scatter gating, combined with 2 additional backbone markers the panel is designed in two consecutive steps. In the first step, a single standardized 10-color-“screening tube” (FITC-HLA-DR, PE-CD15/CD56, ECD-CD5, PC5.5-CD33, PC7-CD13, APC-CD117, APC A700-CD34, APC A750-CD19, PB-CD3, KrO-CD45) is applied for initial orientation of specific lineage assignment. Based on results obtained with the screening tube, a specific multi-tube “classification panel” is used to complete detailed characterization of lineage specific malignancy and maturation stage. Suitable specimens include fresh blood, bone marrow and all body fluids. All samples are stained directly with monoclonal antibodies, followed by the lyses of erythrocytes and a short wash. Compared to standard five color panel previously used the application of greater numbers of informative antibodies in the screening tube and in the 2ndstep muti-tube classification panel is cost and time efficient and results in a more precise characterization of any single event. Conclusion:Our panel construction and algorithm definition for infant and pediatric leukemia immunophenotyping is one of the first 10-color flow cytometry panels described for this application. Advantages are the possibility to obtain highly specific information from minimal specimens with significantly improved laboratory efficiency. The overall performance is currently tested in a routine clinical setting. Disclosures:No relevant conflicts of interest to declare.

Design and validation of bending test method for characterization of miniature pediatric cortical bone specimens

Osteogenesis imperfecta is a genetic disorder of bone fragility; however, the effects of this disorder on bone material properties are not well understood. No study has yet measured bone material strength in humans with osteogenesis imperfecta. Small bone specimens are often extracted during routine fracture surgeries in children with osteogenesis imperfecta. These specimens could provide valuable insight into the effects of osteogenesis imperfecta on bone material strength; however, their small size poses a challenge to their mechanical characterization. In this study, a validated miniature three-point bending test is described that enables measurement of the flexural material properties of pediatric cortical osteotomy specimens as small as 5 mm in length. This method was validated extensively using bovine bone, and the effect of span/depth aspect ratio (5 vs 6) on the measured flexural properties was examined. The method provided reasonable results for both Young's modulus and flexural strength in bovine bone. With a span/depth ratio of 6, the median longitudinal modulus and flexural strength results were 16.1 (range: 14.4-19.3)GPa and 251 (range: 219-293)MPa, respectively. Finally, the pilot results from two osteotomy specimens from children with osteogenesis imperfecta are presented. These results provide the first measures of bone material strength in this patient population.

Development and validation of a serum total testosterone liquid chromatography–tandem mass spectrometry (LC–MS/MS) assay calibrated to NIST SRM 971

BackgroundAt our institution, serum testosterone in adult males is measured by immunoassay while female and pediatric specimens are sent to a reference laboratory for liquid chromatography–tandem mass spectrometry (LC–MS/MS) analysis due to low concentrations. As this is of significant cost, a testosterone LC–MS/MS assay was developed in-house. MethodsA 5500 QTRAP® using electrospray ionization and a Shimadzu Prominence with a C18 column were used. Gradient elution with formic acid, water and methanol:acetonitrile at 0.5ml/min had a 7-min run-time. A liquid–liquid extraction with hexane:ethyl acetate was carried out on 200μl of serum. Multiple reaction monitoring was employed. ResultsSample preparation took ~80min for 21 samples. Six calibrators were used (0–1263ng/dl; concentration assigned by NIST SRM 971) with 3 quality controls (9, 168 and 532ng/dl). The limits of detection and quantitation were 1 and 2ng/dl respectively. Extraction recovery was ~90% and ion suppression ~5%. Within-run and total precision studies yielded <15% CV at the limit of quantitation and <7% CV through the rest of the linear range. Isobaric interferences were baseline separated from testosterone. Method comparisons between this assay, an immunoassay, and another LC–MS/MS assay were completed. ConclusionsAn accurate and sensitive LC–MS/MS assay for total testosterone was developed. Bringing this assay in-house reduces turnaround time for clinicians and patients and saves our institution funds.

Do all Pediatric Urine Specimens Need to Go to the Laboratory?

Objective: This study aimed to evaluate the accuracy of the urine dipstick in diagnosing UTIs in children at a tertiary care centre in Pakistan. Methods: 72 inpatients at the Aga Khan Hospital pediatric ward, getting laboratory urinalysis due to UTI suspicion, were included. Dipstick tests were done on the urine samples being sent to the lab for microscopy. The sensitivity, specificity and likelihood ratios (LRs) of dipstick LE, and pyuria and bacteriuria on microscopy were calculated and compared, using urine culture results as the gold standard for diagnosis. Results: The specificity of dipstick LE, pyuria and bacteriuria were 77%, 77% and 90% respectively, while the positive likelihood ratios were was 28%, 44% and 49% respectively. Urine cultures were done for 58 patients, with 5 positive cultures, so plausible estimates of sensitivity were not made. Conclusions: Urine microscopy is a more accurate screening test for ruling in UTIs than the dipstick. Keeping in mind its diagnostic limitations, the dipstick can be used to help rule in a UTI, although confirmation by cultures is recommended. Further studies are needed to validate these results in children and to evaluate the dipstick’s sensitivity for ruling out disease.

Paediatric salivary gland pathology

This review presents the authors' 17-year experience with 213 paediatric salivary gland specimens from Children's Hospital surgical pathology service. Of 173 non-tumours, 143 were cysts (137 mucocoeles), 26 were inflammatory lesions, 40 were tumours, with 4 miscellaneous lesions. Of the 36 benign tumours, the most frequent were 17 pleomorphic adenomas and 12 lymphangiomas. Of the four malignant tumours, only one was epithelial in origin, acinic cell carcinoma, with one neuroblastoma, lymphoma, and alveolar rhabdomyosarcoma. Literature review supports that most diseases of adult salivary glands can occur in the paediatric age range.

Comparison of the GenMark Diagnostics eSensor Respiratory Viral Panel to Real-Time PCR for Detection of Respiratory Viruses in Children

A novel eSensor respiratory viral panel (eSensor RVP) multiplexed nucleic acid amplification test (GenMark Diagnostics, Inc., Carlsbad, CA) was compared to laboratory-developed real-time PCR assays for the detection of various respiratory viruses. A total of 250 frozen archived pediatric respiratory specimens previously characterized as either negative or positive for one or more viruses by real-time PCR were examined using the eSensor RVP. Overall agreement between the eSensor RVP and corresponding real-time PCR assays for shared analytes was 99.2% (kappa = 0.96 [95% confidence interval {CI}, 0.94 to 0.98]). The combined positive percent agreement was 95.4% (95% CI, 92.5 to 97.3); the negative percent agreement was 99.7% (95% CI, 99.4 to 99.8). The mean real-time PCR threshold cycle (C(T)) value for specimens with discordant results was 39.73 (95% CI, 38.03 to 41.43). Detection of coinfections and correct identification of influenza A virus subtypes were comparable between methods. Of note, the eSensor RVP rhinovirus assay was found to be more sensitive and specific than the corresponding rhinovirus real-time PCR. In contrast, the eSensor RVP adenovirus B, C, and E assays demonstrated some cross-reactivity when tested against known adenovirus serotypes representing groups A through F. The eSensor RVP is robust and relatively easy to perform, it involves a unique biosensor technology for target detection, and its multiplexed design allows for efficient and simultaneous interrogation of a single specimen for multiple viruses. Potential drawbacks include a slower turnaround time and the need to manipulate amplified product during the protocol, increasing the possibility of contamination.

Long-term stability of biochemical markers in pediatric serum specimens stored at −80°C: A CALIPER Substudy

Pediatric serum samples collected from healthy children in the CALIPER (Canadian Laboratory Initiative in Pediatric Reference Interval) project are stored at -80 °C for various periods of time. This study aimed to determine the stability of chemistry, protein, and hormone analytes under these conditions. Serum samples collected from children of 0-18 years of age attending outpatient clinics were pooled into a single pool or into age-group specific pools. Following baseline measurement, each pool was aliquoted and kept frozen at -80 °C until analysis. Samples were analyzed for 57 biochemical markers at monthly intervals over a 10-13 month period and each aliquot was subject to one freeze-thaw cycle before analysis. The analysis was performed on VITROS Chemistry System, COBAS INTEGRA 400 Plus and IMMULITE 2500. Values obtained at monthly intervals were compared to baseline measurements and examined for trends over time. A majority of analytes measured in this study showed no significant time-dependent change relative to baseline or trend over time after up to 13 months of storage. PTH showed up to 27.2% decline after 10 months of storage with most of the decline evident after 2 months. Most analytes showed variability over time, which is thought to reflect assay variability rather than changes in analyte stability. The study shows stability for a majority of analytes stored in serum at -80 °C after up to 13 months of storage. Samples do not require immediate testing for reference interval determination for the selected analytes with possible exception of PTH.

Detection and quantification of influenza C virus in pediatric respiratory specimens by real-time PCR and comparison with infectious viral counts

BackgroundThe epidemiological and clinical impacts of influenza C virus infection may have been underestimated by conventional viral culture screening alone. ObjectiveTo evaluate a newly developed real-time polymerase chain reaction (PCR) assay as a tool for diagnosing influenza C virus infection. Study designThe primers and probe for real-time PCR were designed to amplify the conserved region of the nucleoprotein gene based on the aligned sequences of nine isolates from 1967 to 2010. Respiratory specimens from children collected between January 2010 and August 2010 were examined for the presence of influenza C virus by cell culture and real-time PCR. Specimens that were positive for the virus using real-time PCR were further examined using an infectivity assay with embryonated hen's eggs. ResultsOf the 1203 specimens examined, 34 (2.8%) tested positive for the influenza C virus by cell culture and 51 (4.2%) tested positive by real-time PCR. The mean viral load and infectivity titer in specimens that tested positive using cell culture were 3.97×108copies/ml and 5.43×105EID50/ml, respectively, and those in specimens that were negative using cell culture were 2.18×106copies/ml and 3.67×102EID50/ml, respectively. In the clinical specimens with viral loads less than 105copies/ml, it was not possible to isolate the virus using embryonated hen's eggs. The copy number-to-EID50 ratio of the clinical specimens was much higher, ranging from 32 to 278,000, than those of culture fluid, ranging from 2.3 to 13.5. ConclusionThe real-time PCR assay described here can be used as a sensitive method for diagnosing influenza C virus infection.

The mRNA Expression of Various Angiogenesis-Related Genes in Pediatric Sarcomas and Nonmalignant Lesions of Tissue

For better understanding cancer pathogenesis and searching a potential target for antineoplastic therapy, the authors have studied mRNA expression profile in tissues from 39 children with histological confirmed malignant sarcomas and from 23 patients with bone and soft tissue nonmalignant lesions. mRNA levels of Angiogenesis-related genes VEGFA (including isoforms of 121, 165, 189), VEGFC, VEGFR-1, VEGFR-2, VEGFR-3, HIF-1α, TF, TFPI-1, TFPI-2, uPA, PAI-1 in pediatric specimens were examined using quantitative reverse transcriptase–polymerase chain reaction (qRT-PCR). uPA, HIF-1α, VEGFR-1, VEGFR-2, VEGFR-3, and VEGFC mRNA levels from nonmalignant tissue were significantly higher than those from cancer tissue. On the other hand, isoform VEGFA121 and VEGFA165 and ratio VEGFA165/189 mRNA levels in cancer were higher in comparison with nonmalignant tissue. There was a strong correlation between VEGFA165 and VEGFA189 mRNA expression levels both in cancer tissue and in nonmalignant tissue. In grade 4 tumors in comparison with grade 2 tumors, there was a reduced VEGFA165/189 ratio. Moreover, TFPI-1 and TFPI-2 mRNA levels were significantly lower in sarcomas than in nonmalignant lesions and TFPI-2 was significantly lower in grade 4 tumors than in grade 2. The present data suggested that mRNA overexpression of angiogenesis-related genes is not a prerogative of malignant tissues. The authors supposed that in pediatric bone and soft tissue pathology, high expression of mRNAs of some angiogenesis-related genes may be associated with inflammation and physiological angiogenesis rather than with the development of a malignant tumor. The authors showed the importance of VEGFA121 and/or VEGFA165 and VEGFA165/189 isoform ratio in pediatric sarcomas neoangiogenesis and TFPI-2 for tumors grade 4.

Clinical validation of Xpert MTB/RIF for the diagnosis of extrapulmonary tuberculosis

Extrapulmonary tuberculosis (EPTB) accounts for more than 20% of tuberculosis (TB) cases. Xpert MTB/RIF (Xpert) (Cepheid, Sunnyvale, CA, USA) is a fully automated amplification system, for which excellent results in the diagnosis of pulmonary TB in highly endemic countries have been recently reported. We aimed to assess the performance of the Xpert system in diagnosing EPTB in a low incidence setting. We investigated with Xpert a large number of consecutive extrapulmonary clinical specimens (1,476, corresponding to 1,068 patients) including both paediatric (494) and adult samples. We found, in comparison with a reference standard consisting of combination of culture and clinical diagnosis of TB, an overall sensitivity and specificity of 81.3% and 99.8% for Xpert, while the sensitivity of microscopy was 48%. For biopsies, urines, pus and cerebrospinal fluids the sensitivity exceeded 85%, while it was slightly under 80% for gastric aspirates. It was, in contrast, lower than 50% for cavitary fluids. High sensitivity and specificity (86.9% and 99.7%, respectively) were also obtained for paediatric specimens. Although the role of culture remains central in the microbiological diagnosis of EPTB, the sensitivity of Xpert in rapidly diagnosing the disease makes it a much better choice compared to smear microscopy. The ability to rule out the disease still remains suboptimal.

Fallopian Tube Correlates of Ovarian Serous Borderline Tumors

Ovarian serous borderline tumors (SBTs) are presumed to originate in the ovarian cortex or peritoneal surface. The pathogenetic role of the fallopian tube (FT) is unclear; however, recently, secretory cell outgrowths (SCOUTs) lacking PAX2 expression were described in benign FTs. This study addressed (1) the differentiation characteristics of SBTs and (2) the frequency of SCOUTs lacking PAX2 expression in the FTs of patients with SBTs and compared (3) SCOUT morphology and (4) PAX2 expression with SBTs. SBTs and FT epithelium shared both ciliated (p73) and secretory (HMFG2) differentiation. PAX2-null SCOUT frequency in FT cross-sections from patients with SBTs was 0.28 (110 of 398) versus 0.112 in benign hysterectomies and nearly 0 in pediatric and postpartum sterilization specimens (P = < 0.001). When adjusted for age, the differences narrowed but remained significant (P = 0.010). SCOUTs were heterogeneous, some displaying ciliated differentiation and papillary architecture. Two cases of discrete multifocal papillary SCOUTs in the FTs were associated with SBTs. All SBTs had heterogeneous PAX2 staining with areas of PAX2 loss. This study shows for the first time that PAX2-null SCOUTs are more common in the oviducts of women with SBTs and that loss of PAX2 expression occurs in most SBTs. These discoveries link both morphologic and functional gene (PAX2) alterations in the oviduct to SBTs, similar to that reported in high-grade serous carcinoma. Further study is warranted to clarify the relationship of the oviduct to serous neoplasia.

Comparison of the Idaho Technology FilmArray System to Real-Time PCR for Detection of Respiratory Pathogens in Children

The FilmArray Respiratory Panel (RP) multiplexed nucleic acid amplification test (Idaho Technology, Inc., Salt Lake City, UT) was compared to laboratory-developed real-time PCR assays for the detection of various respiratory viruses and certain bacterial pathogens. A total of 215 frozen archived pediatric respiratory specimens previously characterized as either negative or positive for one or more pathogens by real-time PCR were examined using the FilmArray RP system. Overall agreement between the FilmArray RP and corresponding real-time PCR assays for shared analytes was 98.6% (kappa = 0.92 [95% confidence interval (CI), 0.89 to 0.94]). The combined positive percent agreement was 89.4% (95% CI, 85.4 to 92.6); the negative percent agreement was 99.6% (95% CI, 99.2 to 99.8). The mean real-time PCR threshold cycle (C(T)) value for specimens with discordant results was 36.46 ± 4.54. Detection of coinfections and correct identification of influenza A virus subtypes were comparable to those of real-time PCR when using the FilmArray RP. The greatest comparative difference in sensitivity was observed for adenovirus; only 11 of 24 (45.8%; 95% CI, 27.9 to 64.9) clinical specimens positive for adenovirus by real-time PCR were also positive by the FilmArray RP. In addition, upon testing 20 characterized adenovirus serotypes prepared at high and low viral loads, the FilmArray RP did not detect serotypes 6 and 41 at either level and failed to detect serotypes 2, 20, 35, and 37 when viral loads were low. The FilmArray RP system is rapid and extremely user-friendly, with results available in just over 1 h with almost no labor involved. Its low throughput is a significant drawback for laboratories receiving large numbers of specimens, as only a single sample can be processed at a time with one instrument.

The TOR Kinase Inhibitor INK128 Is Effective in Pre-B Acute Lymphoblastic Leukemia Models

Abstract 2585Pre-B Acute Lymphoblastic Leukemia (B-ALL) remains a significant cause of morbidity and mortality in both adults and children. Targeting signaling pathways that sustain B-ALL cell survival is a promising strategy to improve treatment outcomes. The target of rapamycin (TOR) is a serine/threonine kinase that integrates signals from oncoproteins and the microenvironment to support ALL cell proliferation and survival. A single TOR enzyme in cells forms two multiprotein complexes, TORC1 and TORC2, with distinct regulation and substrates. The classical TOR inhibitor rapamycin inhibits only some functions of TORC1 and does not acutely inhibit TORC2. A new generation of TOR inhibitors has been developed that inhibit all outputs of both TORC1 and TORC2. These ATP-competitive, TOR kinase inhibitors (TOR-KIs) have improved anti-cancer activity in models of both solid tumors and hematological malignancies. Previously we reported that the TOR-KI compound PP242 is effective in mouse models of Philadelphia Chromosome-positive (Ph+) B-ALL, with minimal toxicity to normal hematopoietic cells or immune function (MR Janes et al., Nat. Med. 2010; 16:205). Here we present promising results on the efficacy of a second generation TOR-KI, INK128, in both Ph+ and non-Ph B-ALL. INK128 is an orally active, highly selective TOR-KI that is currently in phase I clinical trials for solid tumors, multiple myeloma and Waldenstrom's Macroglobulinemia. We find that INK128 has similar biological effects as PP242 on Ph+ B-ALL cells, but these effects are apparent at 5-fold lower concentrations in vitro and 80-fold lower doses in mice. In methylcellulose assays using Ph+ and non-Ph B-ALL specimens, INK128 greatly reduces colony numbers and enhances the efficacy of standard-of-care drugs. Both adult and pediatric non-Ph B-ALL specimens are sensitive to growth suppression by INK128. INK128 also causes apoptosis in B-ALL cells cultured on stromal cells. Importantly, INK128 suppresses human Ph+ B-ALL xenograft expansion in mice at doses that do not impair normal hematopoietic cell proliferation. INK128 is also less immunosuppressive than rapamycin. Together these results provide rationale for testing INK128 and other TOR-KIs in clinical trials of B-ALL therapy. Disclosures:Fruman:Intellikine: Membership on an entity's Board of Directors or advisory committees, Research Funding. Janes:Intellikine: Employment. Martin:Intellikine: Employment. Ren:Intellikine: Employment. Liu:Intellikine: Employment. Rommel:Intellikine: Employment.