Single nucleotide polymorphism array-based signature of low hypodiploidy in acute lymphoblastic leukemia.

Low hypodiploidy (30-39 chromosomes) is one of the most prevalent genetic subtypes among adults with ALL and is associated with a very poor outcome. Low hypodiploid clones can often undergo a chromosomal doubling generating a near-triploid clone (60-78 chromosomes). When cytogenetic techniques detect a near triploid clone, a diagnostic challenge may ensue in differentiating presumed duplicated low hypodiploidy from good risk high hyperdiploid ALL (51-67 chromosomes). We used single-nucleotide polymorphism (SNP) arrays to analyze low hypodiploid/near triploid (HoTr) (n = 48) and high hyperdiploid (HeH) (n = 40) cases. In addition to standard analysis, we derived log2 ratios for entire chromosomes enabling us to analyze the cohort using machine-learning techniques. Low hypodiploid and near triploid cases clustered together and separately from high hyperdiploid samples. Using these approaches, we also identified three cases with 50-60 chromosomes, originally called as HeH, which were, in fact, HoTr and two cases incorrectly called as HoTr. TP53 mutation analysis supported the new classification of all cases tested. Next, we constructed a classification and regression tree model for predicting ploidy status with chromosomes 1, 7, and 14 being the key discriminators. The classifier correctly identified 47/50 (94%) HoTr cases. We validated the classifier using an independent cohort of 44 cases where it correctly called 7/7 (100%) low hypodiploid cases. The results of this study suggest that HoTr is more frequent among older adults with ALL than previously estimated and that SNP array analysis should accompany cytogenetics where possible. The classifier can assist where SNP array patterns are challenging to interpret.

Automated disposable small scale reactor for high throughput bioprocess development: a proof of concept study.

The acceleration of bioprocess development for biologics and vaccines can be enabled by automated high throughput technologies. This will alleviate the significant resource burden from the multi-factorial statistical experimentation required for controlling product quality attributes of complex biologics. Recent technology advances have improved clone evaluation and screening, but have struggled to combine the scale down criteria required for both high cell density cell culture and microbial processes, with sufficient automation and disposable technologies to accelerate process development. This article describes the proof of concept evaluations of an automated disposable small scale reactor for high throughput upstream process development. Characterization studies established the small scale stirred tank disposable 250 mL reactor as similar to those of lab and pilot scale. The reactor generated equivalent process performance for industrial biologics processes for therapeutic protein and monoclonal antibody production using CHO cell culture, Pichia pastoris and E. coli. This included similar growth, cell viability, product titer, and product quality. The technology was shown to be robust across multiple runs and met the requirements for the ability to run high cell density processes (>400 g/L wet cell weight) with exponential feeds and sophisticated event triggered processes. Combining this reactor into an automated array of reactors will ultimately be part of a high throughput process development strategy. This will combine upstream, small scale purification with rapid analytics that will dramatically shorten timelines and costs of developing biological processes.

Dravet syndrome and parent associations: the IDEA League experience with comorbid conditions, mortality, management, adaptation, and grief.

The advent of social networking via the Internet and the commercial availability of tests for SCN1A mutations permitted the rapid development and growth of parent-led associations that provide advocacy and support, as well as promote education and research regarding Dravet syndrome (DS) in the last 10 years. The International Dravet syndrome Epilepsy Action League (IDEA League) is a partnership of parents and professionals united in the purpose of creating greater awareness and understanding of DS. In 2004, parents in the IDEA League support network began to collect data from families about their children with DS in order to investigate observations that, in addition to epilepsy, many of the children seemed to share similar problems. The information gained suggests comorbid conditions and raises many hypotheses for further research. The process has led to more rigorous formal studies and an increased understanding of the clinical spectrum of DS. There is an urgent need for collaborative research, comprehensive care, and professional and family education. Mortality appears high, primarily due to sudden unexplained death in epilepsy (SUDEP) and status epilepticus (SE). Most parents wish direct discussions with their child's physician about mortality. The high risk of death and the many other stresses related to DS result in recurrent grief and loss for patients and families and highlights their need for additional advocacy and support.

A targeted, conventional assay, emergency department HIV testing program integrated with existing clinical procedures.

OBJECTIVE: Various HIV testing models have been described, but widespread implementation has lagged. We describe a clinical HIV testing program notable for its use of conventional (nonrapid) assays, native hospital personnel, and an electronic system to aid targeted patient selection. METHODS: Clinical HIV testing program records and hospital emergency department (ED) and laboratory records were reviewed and linked for the period from January 2007 until November 5, 2008. RESULTS: There were 103,475 visits to the ED, for which 1,258 (1.2%) resulted in HIV testing, and 54 (4.3%) were positive for HIV antibody. Result notification was successful for 52 (96%) individuals with positive test results. After reporting to the health department, it was determined that 28 (2.2%) individuals had received a new diagnosis, of whom 89% were linked with care. Mean baseline CD4 counts for new diagnoses for periods 1 through 3, respectively, were (1) unavailable, (2) 138 cells/µL (95% confidence interval [CI] 34 to 242 cells/µL), and (3) 222 cells/µL (95% CI 119 to 325 cells/µL). Overall, mean calculated to be 180 cells/µL (95% CI 16 to 345 cells/µL). CONCLUSION: This ED HIV testing model successfully expanded new patient identification, result notification, and linkage to care. Although this effort falls short of Centers for Disease Control and Prevention recommendations, the model can be implemented widely without external funding for parallel staffing or rapid assays.

Fear of falling among people who have sustained a stroke: a 6-month longitudinal pilot study.

OBJECTIVE: Fear of falling (FoF) after stroke is not well understood. We assessed change in FoF over the first 6 mo after a stroke and compared 6-mo anxiety, depression, balance, and quality of life (QoL) scores between people with and without baseline FoF (at the time of hospital discharge). METHOD: Data for this longitudinal study were collected at baseline and 6 mo. Of the 28 people included at baseline, 18 remained in the study 6 mo later. RESULTS: FoF significantly decreased over time (p = .015). Participants with baseline FoF had higher 6-mo anxiety and depression scores (s = .002 and .005, respectively) and lower QoL scores (p < .001) than did those without baseline FoF. CONCLUSION: The results are suggestive of the need for occupational therapists and their colleagues to consider anxiety and depression variables in managing the needs of poststroke participants experiencing FoF.

Pasture management to minimize the risk of equine laminitis.

The sugar, starch, and fructan content (collectively referred to as nonstructural carbohydrates [NSC]) of pasture plants is dependent on the environmental conditions under which they have grown. Pasture that is stressed by cold, drought, or lack of nutrients can be 2 to 3 times higher in NSC than pasture that grows quickly in warm weather and is adequately watered and fertilized. Horses at risk for laminitis should have access to pasture limited or be removed completely when environmental conditions are conducive to high levels of NSC accumulation.

Ibuprofen and Acetaminophen Versus Intranasal Ketorolac (Sprix) in an Untreated Endodontic Pain Model: A Randomized, Double-blind Investigation.

INTRODUCTION: Previously, ketorolac was available for primary use only via intravenous and intramuscular routes. Its availability in intranasal form offers an alternative route of administration that patients can self-administer. The purpose of this study was to compare the efficacy of intranasal ketorolac (Sprix; Egalet US Inc, Wayne, PA) with a combination of ibuprofen/acetaminophen in an acute pain model of untreated endodontic patients experiencing moderate to severe pain and symptomatic apical periodontitis. METHODS: Seventy patients experiencing moderate to severe pain, a pulpal diagnosis of symptomatic irreversible pulpitis or necrosis, and a periapical diagnosis of symptomatic apical periodontitis participated. Patients were randomly divided into 2 groups and received either 31.5 mg intranasal ketorolac and placebo capsules or 1000 mg acetaminophen/600 mg ibuprofen capsules and a mock nasal spray. Patients recorded perceived pain scores on a visual analog scale every 15 minutes from drug administration up to 240 minutes. The time to 50% pain relief, the first sign of pain relief, and meaningful pain relief were recorded, and the data were analyzed. RESULTS: A decline in reported pain was observed until 120 minutes after dosing, after which reported pain remained relatively constant. There was no significant difference between the 2 groups for the time to 50% pain relief, the first sign of pain relief, or meaningful pain relief. CONCLUSIONS: The effectiveness of intranasal ketorolac was not significantly different from that of a 1000 mg acetaminophen/600 mg ibuprofen combination. Intranasal ketorolac provides a nonnarcotic alternative and an additional route of medication administration to practicing clinicians.

Incidental discovery of an anterior mediastinal angiomyolipoma.

Angiomyolipomas are benign mesenchymal tumors consisting of blood vessels, smooth muscle, and fat. These noninvasive lesions characteristically manifest in renal tissue but may arise less frequently in extrarenal locations including the mediastinum. Only 8 cases of mediastinal angiomyolipomas have been reported in the literature, 3 of which were reported in the English literature, and only 2 of which were located in the anterior mediastinum. We report a surgically confirmed case of an anterior mediastinal angiomyolipoma incidentally discovered in an asymptomatic patient, in whom a screening breast magnetic resonance imaging revealed an incidental 5.7 cm anterior mediastinal mass. Further characterization with contrast-enhanced chest computed tomography confirmed a heterogeneously enhancing anterior mediastinal mass, suggestive of thymoma or lymphoma. The mass was resected and histopathologic evaluation revealed an angiomyolipoma.

A case of mosaic trisomy 19q12-q13.2 with high BMI, macrocephaly, and speech delay: does USF2 determine size in the 19q phenotypes?

Hall et al. (2010) describe a boy with mosaic trisomy of the proximal part of 19q, with obesity, macrocephaly and global developmental delay. The patient is interesting with regard to his cytogenetic abnormality, which is smaller than those previously reported, and does not include the candidate obesity and insulin-resistance genes identified by other authors (Zung et al., 2007; Davidsson et al., 2010) as possible causes of the overweight/obesity seen in four of five previously documented patients. This suggests that a novel obesity locus may reside in the duplicated region 19q13.11­q13.2. We present a phenotypically similar boy with intrachromosomal insertion of material derived from proximal 19q into proximal 19p, causing mosaic trisomy 19q12­q13.2, and consider the role of USF2, a master transcriptional regulator of metabolic genes, in 19q phenotypes.

Nutritional analysis of gastric contents and body condition score at a single time point in feral horses in Australia.

OBJECTIVE: To determine the impact of a free-choice diet on nutritional intake and body condition of feral horses. ANIMALS: Cadavers of 41 feral horses from 5 Australian locations. PROCEDURES: Body condition score (BCS) was determined (scale of 1 to 9), and the stomach was removed from horses during postmortem examination. Stomach contents were analyzed for nutritional variables and macroelement and microelement concentrations. Data were compared among the locations and also compared with recommended daily intakes for horses. RESULTS: Mean BCS varied by location; all horses were judged to be moderately thin. The BCS for males was 1 to 3 points higher than that of females. Amount of protein in the stomach contents varied from 4.3% to 14.9% and was significantly associated with BCS. Amounts of water-soluble carbohydrate and ethanol-soluble carbohydrate in stomach contents of feral horses from all 5 locations were higher than those expected for horses eating high-quality forage. Some macroelement and microelement concentrations were grossly excessive, whereas others were grossly deficient. There was no evidence of ill health among the horses. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that the diet for several populations of feral horses in Australia appeared less than optimal. However, neither low BCS nor trace mineral deficiency appeared to affect survival of the horses. Additional studies on food sources in these regions, including analysis of water-soluble carbohydrate, ethanol-soluble carbohydrate, and mineral concentrations, are warranted to determine the provenance of such rich sources of nutrients. Determination of the optimal diet for horses may need revision.

Optimization of a glycoengineered Pichia pastoris cultivation process for commercial antibody production.

Glycoengineering enabled the production of proteins with human N-linked glycans by Pichia pastoris. This study used a glycoengineered P. pastoris strain which is capable of producing humanized glycoprotein with terminal galactose for monoclonal antibody production. A design of experiments approach was used to optimize the process parameters. Followed by further optimization of the specific methanol feed rate, induction duration, and the initial induction biomass, the resulting process yielded up to 1.6 g/L of monoclonal antibody. This process was also scaled-up to 1,200-L scale, and the process profiles, productivity, and product quality were comparable with 30-L scale. The successful scale-up demonstrated that this glycoengineered P. pastoris fermentation process is a robust and commercially viable process.

Haemopoietic growth factors significantly improve the mitotic index and chromosome quality in cytogenetic cultures of myeloid neoplasia.

Haemopoietic growth factors stimulate bone marrow cell division, differentiation, and survival in vivo. We have investigated the use of recombinant human haemopoietic growth factors in vitro to improve cytogenetic cultures. Using a combination of granulocyte colony stimulating factor, granulocyte macrophage colony stimulating factor, stem cell factor, and interleukin-3, we developed an additive for bone marrow cultures intended to stimulate myeloid cell growth. Sixty-seven paired parallel cultures were analyzed, of which 50 were abnormal. The growth factor (GF) cultures showed a median four- to five-fold increase in mitotic index (MI) (P < 0.0001). In addition, the chromosome morphology was significantly improved in the GF cultures with a median increase in United Kingdom National External Quality Assessment Scheme quality score of 1.25 points (P < 0.0001). There was no statistically significant difference in the number of abnormal cells between the two culture methods. The combination of higher MI and improved chromosome quality substantially reduces the time required to process a case; furthermore, the GF medium is cheaper than the medium with which it was compared. This method is suitable for both diagnostic and follow-up cytogenetic analysis of acute and chronic myeloid neoplasia and is particularly useful for poorly cellular marrow samples or blood samples that would be expected to fail on standard culture. The use of this method has enabled substantial improvements in work efficiency in our oncology cytogenetic laboratory and reduced average reporting times from 9.0 days (2004/5) to 7.1 days (2005/6), despite a 6% increase in sample numbers.

The variant inv(2)(p11.2q13) is a genuinely recurrent rearrangement but displays some breakpoint heterogeneity.

Human chromosome 2 contains large blocks of segmental duplications (SDs), both within and between proximal 2p and proximal 2q, and these may contribute to the frequency of the common variant inversion inv(2)(p11.2q13). Despite their being cytogenetically homogeneous, we have identified four different breakpoint combinations by fluorescence in situ hybridization mapping of 40 cases of inv(2)(p11.2q13) of European origin. For the vast majority of inversions (35/40), the breakpoints fell within the same spanning BACs, which hybridized to both 2p11.2 and 2q13 on the normal and inverted homologues. Sequence analysis revealed that these BACs contain a significant proportion of intrachromosomal SDs with sequence homology to the reciprocal breakpoint region. In contrast, BACs spanning the rare breakpoint combinations contain fewer SDs and with sequence homology only to the same chromosome arm. Using haplotype analysis, we identified a number of related family subgroups with identical or very closely related haplotypes. However, the majority of cases were not related, demonstrating for the first time that the inv(2)(p11.2q13) is a truly recurrent rearrangement. Therefore, there are three explanations to account for the frequent observation of the inv(2)(p11.2q13): the majority have arisen independently in different ancestors, while a minority either have been transmitted from a common founder or have different breakpoints at the molecular cytogenetic level.