The polo-like kinase 1 (PLK1) inhibitor NMS-P937 is effective in a new model of disseminated primary CD56+ acute monoblastic leukaemia

CD56 is expressed in 15–20% of acute myeloid leukaemias (AML) and is associated with extramedullary diffusion, multidrug resistance and poor prognosis. We describe the establishment and characterisation of a novel disseminated model of AML (AML-NS8), generated by injection into mice of leukaemic blasts freshly isolated from a patient with an aggressive CD56+ monoblastic AML (M5a). The model reproduced typical manifestations of this leukaemia, including presence of extramedullary masses and central nervous system involvement, and the original phenotype, karyotype and genotype of leukaemic cells were retained in vivo. Recently Polo-Like Kinase 1 (PLK1) has emerged as a new candidate drug target in AML. We therefore tested our PLK1 inhibitor NMS-P937 in this model either in the engraftment or in the established disease settings. Both schedules showed good efficacy compared to standard therapies, with a significant increase in median survival time (MST) expecially in the established disease setting (MST = 28, 36, 62 days for vehicle, cytarabine and NMS-P937, respectively). Importantly, we could also demonstrate that NMS-P937 induced specific biomarker modulation in extramedullary tissues. This new in vivo model of CD56+ AML that recapitulates the human tumour lends support for the therapeutic use of PLK1 inhibitors in AML

NMDA receptors and BAX are essential for Aβ impairment of LTP

Accumulation of amyloid-β (Aβ) is a hallmark of Alzheimer’s disease, a neurodegenerative disorder in which synapse loss and dysfunction are early features. Acute exposure of hippocampal slices to Aβ leads to changes in synaptic plasticity, specifically reduced long-term potentiation (LTP) and enhanced long-term depression (LTD), with no change in basal synaptic transmission. We also report here that D-AP5, a non-selective NMDA receptor antagonist, completely prevented Aβ-mediated inhibition of LTP in area CA1 of the hippocampus. Ro25-6981, an antagonist selective for GluN2B (NR2B) NMDA receptors, only partially prevented this Aβ action, suggesting that GluN2A and GluN2B receptors may both contribute to Aβ suppression of LTP. The effect of Aβ on LTP was also examined in hippocampal slices from BAX −/− mice and wild-type littermates. Aβ failed to block LTP in hippocampal slices from BAX −/− mice, indicating that BAX is essential for Aβ inhibition of LTP

Immunoglobulin E-binding site in Fc epsilon receptor (Fc epsilon RII/CD23) identified by homolog-scanning mutagenesis

The IgE-binding site of the human low-affinity receptor for IgE (Fc epsilon RII/CD23) has previously been mapped to the extracellular domain between amino acid residues 160 and 287. We now have investigated which conformational epitope within this domain specifies the receptor-ligand interaction. The analysis of homolog-scanning mutants expressed in mammalian cells demonstrates that amino acid side chains that affect IgE binding are located in two discontinuous segments, between residues 165-190 and 224-256. The overall structure of the chimeric binding domains, as probed with 11 conformation-sensitive monoclonal antibodies, is generally not distorted, except by replacement of residues 165-183. In this region, disruption of binding function appears to be caused by global conformational constraints on the binding site. Substitution and deletion mutants demonstrate that six out of eight extracellular cysteines, Cys163, Cys174, Cys191, Cys259, Cys273, and Cys282, are necessary for IgE binding and are most likely involved in intramolecular disulfide bridges. We show that the Fc epsilon RII domain delineated by Cys163 and Cys282 encodes all the structural information required to form the IgE-binding site

Impaired cerebral cortex development and blood pressure regulation in FGF-2-deficient mice.

Fibroblast growth factor-2 (FGF-2) has been implicated in various signaling processes which control embryonic growth and differentiation, adult physiology and pathology. To analyze the in vivo functions of this signaling molecule, the FGF-2 gene was inactivated by homologous recombination in mouse embryonic stem cells. FGF-2-deficient mice are viable, but display cerebral cortex defects at birth. Bromodeoxyuridine pulse labeling of embryos showed that proliferation of neuronal progenitors is normal, whereas a fraction of them fail to colonize their target layers in the cerebral cortex. A corresponding reduction in parvalbumin-positive neurons is observed in adult cortical layers. Neuronal defects are not limited to the cerebral cortex, as ectopic parvalbumin-positive neurons are present in the hippocampal commissure and neuronal deficiencies are observed in the cervical spinal cord. Physiological studies showed that FGF-2-deficient adult mice are hypotensive. They respond normally to angiotensin II-induced hypertension, whereas neural regulation of blood pressure by the baroreceptor reflex is impaired. The present genetic study establishes that FGF-2 participates in controlling fates, migration and differentiation of neuronal cells, whereas it is not essential for their proliferation. The observed autonomic dysfunction in FGF-2-deficient adult mice uncovers more general roles in neural development and function

In vivo imaging of early stage apoptosis by measuring real-time caspase-3/7 activation

In vivo imaging of apoptosis in a preclinical setting in anticancer drug development could provide remarkable advantages in terms of translational medicine. So far, several imaging technologies with different probes have been used to achieve this goal. Here we describe a bioluminescence imaging approach that uses a new formulation of Z-DEVD-aminoluciferin, a caspase 3/7 substrate, to monitor in vivo apoptosis in tumor cells engineered to express luciferase. Upon apoptosis induction, Z-DEVD-aminoluciferin is cleaved by caspase 3/7 releasing aminoluciferin that is now free to react with luciferase generating measurable light. Thus, the activation of caspase 3/7 can be measured by quantifying the bioluminescent signal. Using this approach, we have been able to monitor caspase-3 activation and subsequent apoptosis induction after camptothecin and temozolomide treatment on xenograft mouse models of colon cancer and glioblastoma, respectively. Treated mice showed more than 2-fold induction of Z-DEVD-aminoluciferin luminescent signal when compared to the untreated group. Combining D-luciferin that measures the total tumor burden, with Z-DEVD-aminoluciferin that assesses apoptosis induction via caspase activation, we confirmed that it is possible to follow non-invasively tumor growth inhibition and induction of apoptosis after treatment in the same animal over time. Moreover, here we have proved that following early apoptosis induction by caspase 3 activation is a good biomarker that accurately predicts tumor growth inhibition by anti-cancer drugs in engineered colon cancer and glioblastoma cell lines and in their respective mouse xenograft models

Magnetic resonance imaging and histopathological characterization of prostate tumors in TRAMP mice as model for pre-clinical trials

BACKGROUND: The Transgenic Adenocarcinoma of Mouse Prostate (TRAMP) develops progressive forms of prostate cancer. Due to the lack of a validated non-invasive methodology, pathology has been so far the most common parameter evaluated in efficacy studies. METHODS: We studied by magnetic resonance imaging (MRI) 210 mice that were repeatedly measured up to 33 weeks of age in order to stage prostate tumors and follow pathological progression in single animals. A pre-clinical trial with doxorubicin was also performed. RESULTS: Progressive forms of cancer (well and poorly differentiated (PD) adenocarcinomas) were easily recognized on MR images and MRI findings were validated against histopathological analysis. Age at tumor onset was different for the two tumoral forms. Doxorubicin treatment caused a strong reduction in tumor volume. CONCLUSIONS: Prostate cancer in TRAMP mice is multifocal and heterogeneous: a non-invasive methodology such as MRI facilitates the rational design of translational pre-clinical trials in this widely used animal model