Negative selection of chronic lymphocytic leukaemia cells using a bifunctional rosette-based antibody cocktail

<p>Abstract</p> <p>Background</p> <p>High purity of tumour samples is a necessity for accurate genetic and expression analysis and is usually achieved by positive selection in chronic lymphocytic leukaemia (CLL).</p> <p>Results</p> <p>We adapted a bifunctional rosette-based antibody cocktail for negative selection of B-cells for isolating CLL cells from peripheral blood (PB). PB samples from CLL patients were split into aliquots. One aliquot of each sample was enriched by density gradient centrifugation (DGC), while the other aliquot of each sample was incubated with an antibody cocktail for B-cell enrichment prior to DGC (RS+DGC). The purity of CLL cells after DGC averaged 74.1% (range: 15.9 – 97.4%). Using RS+DGC, the purity averaged 93.8% (range: 80.4 – 99.4%) with 23 of 29 (79%) samples showing CLL purities above 90%. RNA extracted from enriched CLL cells was of appropriately high quality for microarray analysis.</p> <p>Conclusion</p> <p>This study confirms the use of a bifunctional rosette-based antibody cocktail as an effective method for the purification of CLL cells from peripheral blood.</p

The trials and tribulations of conducting an m-health pilot randomized controlled trial to improve oral cancer therapy adherence: recommendations for future multisite, non-drug clinical trials

Abstract Objective Integrating mobile phone-based health (m-health) interventions into healthcare systems is one solution to improve access to services for the growing number of patients with chronic illness. Practical challenges such as poor recruitment and inadequate resource allocation can hamper the assessment of such interventions with clinical trial methodology. This paper highlights the challenges encountered during a pilot randomized controlled trial of an m-health medication adherence intervention and offers recommendations for future multi-site, non-drug clinical trials. Results Eighteen patients were recruited to the study; eight were randomly allocated to the intervention arm. Intervention participants responded to their daily medication-reminder text messages, indicating that medication had been taken or not, and nurses were able to organize their calls around their workload. The trial closed prematurely primarily due to inadequate numbers of eligible patients; however, other potentially resolvable feasibility issues were identified. These included lack of infrastructure at study sites, poor screening data acquisition and management processes, and inexperience in conducting supportive care trials at participating sites. M-health intervention trials are designed to inform implementation of best supportive care practice. Adequate skills and infrastructure are research prerequisites that require careful consideration and sufficient investment for the successful execution of multi-site supportive care trials. Trial registration Australian and New Zealand Clinical Trials Register: ACTRN1261200063586

Detection of NPM1 exon 12 mutations and FLT3 – internal tandem duplications by high resolution melting analysis in normal karyotype acute myeloid leukemia

<p>Abstract</p> <p>Background</p> <p>Molecular characterisation of normal karyotype acute myeloid leukemia (NK-AML) allows prognostic stratification and potentially can alter treatment choices and pathways. Approximately 45–60% of patients with NK-AML carry <it>NPM1 </it>gene mutations and are associated with a favourable clinical outcome when <it>FLT3</it>-internal tandem duplications (ITD) are absent. High resolution melting (HRM) is a novel screening method that enables rapid identification of mutation positive DNA samples.</p> <p>Results</p> <p>We developed HRM assays to detect <it>NPM1 </it>mutations and <it>FLT3</it>-ITD and tested diagnostic samples from 44 NK-AML patients. Eight were <it>NPM1 </it>mutation positive only, 4 were both <it>NPM1 </it>mutation and <it>FLT3</it>-ITD positive and 4 were <it>FLT3</it>-ITD positive only. A novel point mutation Y572C (c.1715A>G) in exon 14 of <it>FLT3 </it>was also detected. In the group with <it>de novo </it>NK-AML, 40% (12/29) were <it>NPM1 </it>mutation positive whereas <it>NPM1 </it>mutations were observed in 20% (3/15) of secondary NK-AML cases. Sequencing was performed and demonstrated 100% concordance with the HRM results.</p> <p>Conclusion</p> <p>HRM is a rapid and efficient method of screening NK-AML samples for both novel and known <it>NPM1 </it>and <it>FLT3 </it>mutations. <it>NPM1 </it>mutations can be observed in both primary and secondary NK-AML cases.</p

Nanomedicines for the delivery of biologics

A special symposium of the Academy of Pharmaceutical Sciences Nanomedicines Focus Group reviewed the current status of the use of nanomedicines for the delivery of biologics drugs. This meeting was particularly timely with the recent approval of the first siRNA-containing product Onpattro™ (patisiran), which is formulated as a lipid nanoparticle for intravenous infusion, and the increasing interest in the use of nanomedicines for the oral delivery of biologics. The challenges in delivering such molecules were discussed with specific emphasis on the delivery both across and into cells. The latest developments in Molecular Envelope Technology® (Nanomerics Ltd, London, UK), liposomal drug delivery (both from an academic and industrial perspective), opportunities offered by the endocytic pathway, delivery using genetically engineered viral vectors (PsiOxus Technologies Ltd, Abingdon, UK), Transint™ technology (Applied Molecular Transport Inc., South San Francisco, CA, USA), which has the potential to deliver a wide range of macromolecules, and AstraZeneca’s initiatives in mRNA delivery were covered with a focus on their uses in difficult to treat diseases, including cancers. Preclinical data were presented for each of the technologies and where sufficiently advanced, plans for clinical studies as well as early clinical data. The meeting covered the work in progress in this exciting area and highlighted some key technologies to look out for in the future

Immunochemotherapy With Obinutuzumab or Rituximab for Previously Untreated Follicular Lymphoma in the GALLIUM Study: Influence of Chemotherapy on Efficacy and Safety

PurposeThe GALLIUM study (ClinicalTrials.gov identifier: NCT01332968) showed that obinutuzumab (GA101;G) significantly prolonged progression-free survival (PFS) in previously untreated patients with follicular lymphoma relative to rituximab (R) when combined with cyclophosphamide (C), doxorubicin, vincristine (V), and prednisone (P;CHOP);CVP;or bendamustine. This report focuses on the impact of chemotherapy backbone on efficacy and safety.Patients and Methods: A total of 1,202 patients with previously untreated follicular lymphoma (grades 1 to 3a), advanced disease (stage III or IV, or stage II with tumor diameter 7 cm), Eastern Cooperative Oncology Group performance status 0 to 2, and requiring treatment were randomly assigned 1:1 to G 1,000 mg on days 1, 8, and 15 of cycle 1 and day 1 of subsequent cycles or R 375 mg/m(2) on day 1 of each cycle, for six to eight cycles, depending on chemotherapy (allocated nonrandomly by center). Responding patients received G or R for 2 years or until disease progression.Results: Baseline Follicular Lymphoma International Prognostic Index risk, bulky disease, and comorbidities differed by chemotherapy. After 41.1 months median follow-up, PFS (primary end point) was superior for G plus chemotherapy (overall hazard ratio [HR], 0.68;95% CI, 0.54 to 0.87;P = .0016), with consistent results across chemotherapy backbones (bendamustine: HR, 0.63;95% CI, 0.46 to 0.88;CHOP: HR, 0.72;95% CI, 0.48 to 1.10;CVP: HR, 0.79;95% CI, 0.42 to 1.47). Grade 3 to 5 adverse events, notably cytopenias, were most frequent with CHOP. Grade 3 to 5 infections and second neoplasms were most frequent with bendamustine, which was associated with marked and prolonged reductions in T-cell counts. Fatal events were more frequent in patients treated with bendamustine, possibly reflecting differences in patient risk profiles.Conclusion: Improved PFS was observed for G plus chemotherapy for all three chemotherapy backbones. Safety profiles differed, although comparisons are confounded by nonrandom chemotherapy allocation

Chemistry courses as the turning point for premedical students

Previous research has documented that negative experiences in chemistry courses are a major factor that discourages many students from continuing in premedical studies. This adverse impact affects women and students from under-represented minority (URM) groups disproportionately. To determine if chemistry courses have a similar effect at a large public university, we surveyed 1,036 students from three entering cohorts at the University of California, Berkeley. We surveyed students at the beginning of their first year at the university and again at the end of their second year. All subjects had indicated an interest in premedical studies at the time they entered the university. We conducted follow-up interviews with a stratified sub-set of 63 survey respondents to explore the factors that affected their level of interest in premedical studies. Using a 10-point scale, we found that the strength of interest in premedical studies declined for all racial/ethnic groups. In the follow-up interviews, students identified chemistry courses as the principal factor contributing to their reported loss of interest. URM students especially often stated that chemistry courses caused them to abandon their hopes of becoming a physician. Consistent with reports over more than 50 years, it appears that undergraduate courses in chemistry have the effect of discouraging otherwise qualified students, as reflected in their admission to one of the most highly selective public universities in the US, from continuing in premedical studies, especially in the case of URM students. Reassessment of this role for chemistry courses may be overdue