Expression of a homodimeric type I cytokine receptor is required for JAK2V617F-mediated transformation

A recurrent somatic activating mutation in the nonreceptor tyrosine kinase JAK2 (JAK2V617F) occurs in the majority of patients with the myeloproliferative disorders polycythemia vera, essential thrombocythemia, myelofibrosis with myeloid metaplasia, and, less commonly, chronic myelomonocytic leukemia. We do not understand the basis for the specificity of the JAK2V617F mutation in clonal disorders of the myeloid, but not lymphoid, lineage, nor has the basis for the pleiotropic phenotype of JAK2V617F-associated myeloproliferative disorders been delineated. However, the presence of the identical mutation in patients with related, but clinicopathologically distinct, myeloid disorders suggests that interactions between the JAK2V617F kinase and other signaling molecules may influence the phenotype of hematopoietic progenitors expressing JAK2V617F. Here, we show that coexpression of the JAK2V617F mutant kinase with a homodimeric Type I cytokine receptor, the erythropoietin receptor (EpoR), the thrombopoietin receptor, or the granulocyte colony-stimulating-factor receptor, is necessary for transformation of hematopoietic cells to growth-factor independence and for hormone-independent activation of JAK-STAT signaling. Furthermore, EpoR mutations that impair erythropoietin-mediated JAK2 or STAT5 activation also impair transformation mediated by the JAK2V617F kinase, indicating that JAK2V617F requires a cytokine receptor scaffold for its transforming and signaling activities. Our results reveal the molecular basis for the prevalence of JAK2V617F in diseases of myeloid lineage cells that express these Type I cytokine receptors but not in lymphoid lineage cells that do not.     

CD34(+)/CD38(-) stem cells in chronic myeloid leukemia express Siglec-3 (CD33) and are responsive to the CD33-targeting drug gemtuzumab/ozogamicin

Background

CD33 is a well-known stem cell target in acute myeloid leukemia. So far, however, little is known about expression of CD33 on leukemic stem cells in chronic leukemias.

Design and Methods

We analyzed expression of CD33 in leukemic progenitors in chronic myeloid leukemia by multi-color flow cytometry and quantitative polymerase chain reaction. In addition, the effects of a CD33-targeting drug, gemtuzumab/ozogamicin, were examined.

Results

As assessed by flow cytometry, stem cell-enriched CD34⁺/CD38⁻/CD123⁺ leukemic cells expressed significantly higher levels of CD33 compared to normal CD34⁺/CD38⁻ stem cells. Moreover, highly enriched leukemic CD34⁺/CD38⁻ cells (>98% purity) displayed higher levels of CD33 mRNA. In chronic phase patients, CD33 was found to be expressed invariably on most or all stem cells, whereas in accelerated or blast phase of the disease, the levels of CD33 on stem cells varied from donor to donor. The MDR1 antigen, supposedly involved in resistance against ozogamicin, was not detectable on leukemic CD34⁺/CD38⁻ cells. Correspondingly, gemtuzumab/ozogamicin produced growth inhibition in leukemic progenitor cells in all patients tested. The effects of gemtuzumab/ozogamicin were dose-dependent, occurred at low concentrations, and were accompanied by apoptosis in suspension culture. Moreover, the drug was found to inhibit growth of leukemic cells in a colony assay and long-term culture-initiating cell assay. Finally, gemtuzumab/ozogamicin was found to synergize with nilotinib and bosutinib in inducing growth inhibition in leukemic cells.

Conclusions

CD33 is expressed abundantly on immature CD34⁺/CD38⁻ stem cells and may serve as a stem cell target in chronic myeloid leukemia.     

Fibroblast growth factor receptors as therapeutic targets in human melanoma: synergism with BRAF inhibition

Cutaneous melanoma is a tumor with rising incidence and a very poor prognosis at the disseminated stage. Melanomas are characterized by frequent mutations in BRAF and also by overexpression of fibroblast growth factor 2 (FGF2), offering opportunities for therapeutic intervention. We investigated inhibition of FGF signaling and its combination with dacarbazine or BRAF inhibitors as an antitumor strategy in melanoma. The majority of melanoma cell lines displayed overexpression of FGF2 but also FGF5 and FGF18 together with different isoforms of FGF receptors (FGFRs) 1-4. Blockade of FGF signals with dominant-negative receptor constructs (dnFGFR1, 3, or 4) or small-molecule inhibitors (SU5402 and PD166866) reduced melanoma cell proliferation, colony formation, as well as anchorage-independent growth, and increased apoptosis. DnFGFR constructs also significantly inhibited tumor growth in vivo. Combination of FGF inhibitors with dacarbazine showed additive or antagonistic effects, whereas synergistic drug interaction was observed when combining FGFR inhibition with the multikinase/BRAF inhibitor sorafenib or the V600E mutant-specific BRAF inhibitor RG7204. In conclusion, FGFR inhibition has antitumor effects against melanoma cells in vitro and in vivo. Combination with BRAF inhibition offers a potential for synergistic antimelanoma effects and represents a promising therapeutic strategy against advanced melanoma.     

The role of dendritic cells in alphaherpesvirus infections: archetypes and paradigms

Dendritic cells (DCs) play a critical role in orchestrating both innate and adaptive components of the immune system and are therefore of pivotal importance in the initiation of immune responses to control and eliminate viral infections. A major focus of this review is to give an overview on the recent findings that point out the importance of DCs in controlling alphaherpesvirus infections, but also indicate that these viruses have evolved several strategies to inhibit and/or exploit DC functions to delay or escape elimination by the immune system. In addition, we point out the common features and interspecies differences between DCs from man and animal, and discuss the potential use of animal alphaherpesvirus homologues to gain further insights into the interaction between alphaherpesviruses and DCs in their natural virus–host environment. Finally, recent knowledge on the potential of alphaherpesviruses as vectors for DC stimulation and their use for immunotherapy is presented. Copyright © 2009 John Wiley & Sons, Ltd.     

Hair trace elementary profiles in aging rodents and primates: links to altered cell homeodynamics and disease

Aging is associated with an increased incidence of pathological conditions such as neurodegeneration, cardiovascular and renal disease, and cancer. These conditions are believed to be linked to a disruption in cell homeodynamics, which is regulated by essential trace elements. In this study we used hair elementary analysis by inductively coupled plasma mass spectrometry (ICPMS) to examine age-related profiles of 47 elements in both rats and common marmoset monkeys. Hair was collected from young adult (6 months) and aged (18 months) Long–Evans male rats, and young adult (2 years), middle-aged (4 years) and aged (>8 years) marmosets. The results revealed that aging reduces content levels of cobalt, potassium and selenium while content levels of aluminium, arsenic, boron, mercury, molybdenum, and titanium were elevated in aged rats. Similarly, aged marmosets showed reduced levels of cobalt and elevated levels of aluminium. Case studies in aged rats revealed that myocardial infarction was associated with elevated levels of sodium, potassium and cadmium and reduced zinc, while renal failure was linked to elevated content of potassium, chloride and boron and reduced contents of manganese. Carcinoma was linked to elevated arsenic and reduced selenium levels. These findings indicate that hair elementary profiles in healthy aging and age-related diseases reflect altered cell and organ metabolic functions. Cobalt and aluminium in particular may serve as biomarkers of aging in animal models. Thus, elementary deposition in hair may have predictive and diagnostic value in age-related pathological conditions, including cardiovascular and kidney disease and cancer.     

Characterization of spatial performance in male and female Long-Evans rats by means of the Morris water task and the ziggurat task

Sex differences are prominent influences on spatial performance. One of the most common tasks to assess sex differences in spatial navigation in rodents is the Morris water task (MWT). In this task rats swim in a pool of water to locate a hidden platform employing the topographical relationships among the distal visual cues, pool wall, and goal location. Some evidence suggests that male rats display superior performance relative to females in the MWT. It is unknown, however, to what extent the sex difference in rats is task-dependent. This study compared the performance of male and female Long-Evans rats in the wet-land MWT versus the dry-land ziggurat task (ZT). The ZT represents a new dry-land task in which rats explore an arena with 16 ziggurat pyramids to locate food rewards. Several behavioural parameters, including latency, path length, path speed, probe trial performance, errors, and the number of returns were used as indices of spatial learning and memory. While males and females did not display significant differences in the traditional measures of spatial navigation within MWT, they displayed a robust sex difference in all measures of the ZT. These results indicate task-specific sex differences in spatial performance. Our findings suggest that males and females may employ different learning strategies in the MWT and ZT and that the latter task provides a more favourable task for assessing sex differences in rats.