Ex vivo Drug Sensitivity Imaging-based Platform for Primary Acute Lymphoblastic Leukemia Cells.

Resistance of acute lymphoblastic leukemia (ALL) cells to chemotherapy, whether present at diagnosis or acquired during treatment, is a major cause of treatment failure. Primary ALL cells are accessible for drug sensitivity testing at the time of new diagnosis or at relapse, but there are major limitations with current methods for determining drug sensitivity ex vivo. Here, we describe a functional precision medicine method using a fluorescence imaging platform to test drug sensitivity profiles of primary ALL cells. Leukemia cells are co-cultured with mesenchymal stromal cells and tested with a panel of 40 anti-leukemia drugs to determine individual patterns of drug resistance and sensitivity ("pharmacotype"). This imaging-based pharmacotyping assay addresses the limitations of prior ex vivo drug sensitivity methods by automating data analysis to produce high-throughput data while requiring fewer cells and significantly decreasing the labor-intensive time required to conduct the assay. The integration of drug sensitivity data with genomic profiling provides a basis for rational genomics-guided precision medicine. Key features Analysis of primary acute lymphoblastic leukemia (ALL) blasts obtained at diagnosis from bone marrow aspirate or peripheral blood. Experiments are performed ex vivo with mesenchymal stromal cell co-culture and require four days to complete. This fluorescence imaging-based protocol enhances previous ex vivo drug sensitivity assays and improves efficiency by requiring fewer primary cells while increasing the number of drugs tested to 40. It takes approximately 2-3 h for sample preparation and processing and a 1.5-hour imaging time. Graphical overview.

Epigenetic activation of the FLT3 gene by ZNF384 fusion confers a therapeutic susceptibility in acute lymphoblastic leukemia.

FLT3 is an attractive therapeutic target in acute lymphoblastic leukemia (ALL) but the mechanism for its activation in this cancer is incompletely understood. Profiling global gene expression in large ALL cohorts, we identify over-expression of FLT3 in ZNF384-rearranged ALL, consistently across cases harboring different fusion partners with ZNF384. Mechanistically, we discover an intergenic enhancer element at the FLT3 locus that is exclusively activated in ZNF384-rearranged ALL, with the enhancer-promoter looping directly mediated by the fusion protein. There is also a global enrichment of active enhancers within ZNF384 binding sites across the genome in ZNF384-rearranged ALL cells. Downregulation of ZNF384 blunts FLT3 activation and decreases ALL cell sensitivity to FLT3 inhibitor gilteritinib in vitro. In patient-derived xenograft models of ZNF384-rearranged ALL, gilteritinib exhibits significant anti-leukemia efficacy as a monotherapy in vivo. Collectively, our results provide insights into FLT3 regulation in ALL and point to potential genomics-guided targeted therapy for this patient population.

Preclinical evaluation of proteolytic targeting of LCK as a therapeutic approach in T cell acute lymphoblastic leukemia.

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy, and there is an unmet need for targeted therapies, especially for patients with relapsed disease. We have recently identified pre-T cell receptor and lymphocyte-specific protein tyrosine kinase (LCK) signaling as a common therapeutic vulnerability in T-ALL. LCK inhibitor dasatinib showed efficacy against T-ALL in preclinical studies and in patients with T-ALL; however, this is transient in most cases. Leveraging the proteolysis targeting chimera (PROTAC) approach, we developed a series of LCK degraders using dasatinib as an LCK ligand and phenyl-glutarimide as a cereblon-directing moiety. Our lead compound SJ11646 exhibited marked efficiency in cereblon-mediated LCK degradation in T-ALL cells. Relative to dasatinib, SJ11646 showed up to three orders of magnitude higher cytotoxicity in LCK-activated T-ALL cell lines and primary leukemia samples in vitro, with drastically prolonged suppression of LCK signaling. In vivo pharmacokinetic and pharmacodynamic profiling indicated a 630% increase in the duration of LCK suppression by SJ11646 over dasatinib in patient-derived xenograft models of T-ALL, which translated into its extended leukemia-free survival over dasatinib in vivo. Last, SJ11646 retained a high binding affinity to 51 human kinases, particularly ABL1, KIT, and DDR1, all of which are known drug targets in other cancers. Together, our dasatinib-based phenyl-glutarimide PROTACs are promising therapeutic agents in T-ALL and valuable tools for developing degradation-based therapeutics for other cancers.

The potential for sociocultural factors in the diagnosis of ADHD in children.

PURPOSE: The nongenetic contributors to attention deficit/hyperactivity disorder (ADHD) remain to be identified. A previous study in eastern Wisconsin (prevalence 13.5%) suggested that male gender, white race, lower block group median household income and population density, and greater distance to the nearest park were factors predictive of ADHD diagnosis. We performed a similar study in Dane County, Wisconsin. METHODS: Cross sectional study of children age 5-17, with and without ADHD diagnosis, who received well child care in Dane County UW Family Medicine clinics (N=7954) 2007-2008. Street addresses were geocoded to 2000 Census block group. Univariate analysis was done by chi-square test or Mann-Whitney U test, multivariate analysis by logistic regression. RESULTS: ADHD diagnosis was present in 309 (3.9%) children (74.1% male; P = 0.000, compared to females) and more frequently diagnosed in black children (6.8% of black children had ADHD diagnosis) than white (4%), Native American (2.7%), Hispanic (1.6%), or Asian (1.3%) children. In contrast to eastern Wisconsin and to Milwaukee County (a subset of the eastern Wisconsin study where black rates were identical to that of Dane County), black race rather than white race was predictive of ADHD in Dane County, while median household income, population density, and distance to nearest park were not associated. The range of ADHD within school district boundaries was 2.4%-7.1% (for N > 100/district). In the group of districts with >4% ADHD diagnosis, the increased rates were largely among whites. CONCLUSION: ADHD diagnosis was much less common in this Dane County cohort than in eastern Wisconsin and was more common among blacks, but not predicted by other geo-demographic factors. Like eastern Wisconsin, ADHD diagnosis prevalence varied with apparent school district boundaries.

Púrpura palpable asociada con hepatitis C / Papable purpura related to hepatitis C

Entre las múltiples manifestaciones clínicas extrahepáticas asociadas con la hepatitis C descritas hasta el momento se encuentran las dermatológicas, síntomas que pueden ser de gran ayuda en el diagnóstico temprano de la enfermedad y, por ende, en la disminución de su morbilidad y mortalidad. La púrpura palpable es una de ellas. A continuación describimos el caso de una mujer con cirrosis hépatica asociada con púrpura palpable y hepatitis C