BACKGROUND:: Healthy children presenting with neutropenia are often hospitalized and treated empirically with antibiotics without an evidence of infection. The objective of this study was to investigate the infectious causes of isolated transient neutropenia in otherwise previously- healthy children. METHOD:: A two-year prospective study was conducted at a tertiary hospital in Kuwait. All previously-healthy children (aged 1 month-12 years) hospitalized with isolated neutropenia defined as absolute neutrophil count (ANC) ≤ 1.5x 10/L were enrolled in the study. Investigations to identify the infectious causes included blood and urine culture for bacteria whereas for viruses, serology for EBV, CMV, adenovirus, parvovirus and PCR for HHV6 and enterovirus were performed. RESULTS:: Fifty five children were enrolled during the study. Children less than 2 years constituted 73% of the sample. There were two peaks of presentation: March-May (33%) and September- November (38%). Associated features were: congested throat (56%), runny nose (53%), and cervical lymphadenopathy (20%). The median ANC on admission was 0.6 x 10/L. Associated infections were documented in 55% of enrolled children and were as follows: HHV6 30%, enterovirus 23%, influenza A H1N113%, parvovirus10%, EBV10%, UTI (E.coli) 7%, and adenovirus 7%. No serious bacterial infection was identified and the mean time for recovery of the ANC was 16.7 ± 15 days. CONCLUSION:: Neutropenia in previously-healthy children in Kuwait is caused by demonstrable infections in 55% of cases. Majority of children will recover their ANC completely within one month without significant infectious complications.

The spread of multiple myeloma (MM) involves (re)circulation into the peripheral blood and (re)entrance or homing of MM cells into new sites of the BM. Hypoxia in solid tumors was shown to promote metastasis through activation of proteins involved in the endothelial to mesenchymal transition (EMT) process. In this study, we hypothesized that MM associated hypoxic conditions activates EMT related proteins and promotes metastasis of MM cells. Hypoxia activated EMT-related machinery in MM cells, decreased expression of E-cadherin and consequently decreased adhesion of MM cells to the BM and enhanced egress of MM cells to the circulation. In parallel, hypoxia increased the expression of CXCR4, and consequently increased the migration and homing of circulating MM cells to new BM niches. Further studies to manipulate hypoxia in order to regulate tumor dissemination as a therapeutic strategy are warranted.

Drug resistance is a growing area of concern. It has been shown that a small, residual pool of leukemic CD34+ progenitor cells can survive in the marrow microenvironment of chronic myeloid leukemia (CML) patients after years of kinase inhibitor treatment. Bone marrow (BM) stroma has been implicated in the long-term survival of leukemic cells, and contributes to the expansion and proliferation of both transformed and normal hematopoietic cells. Mechanistically, we found that CML cells expressed CXCR4, and that plerixafor diminished BCR-ABL-positive cell migration and reduced adhesion of these cells to extra cellular-matrix components and to BM stromal cells in vitro. Moreover, plerixafor decreased the drug resistance of CML cells induced by co-culture with BM stromal cells in vitro. Using a functional mouse model of progressive and residual disease, we demonstrated the ability of the CXCR4 inhibitor, plerixafor, to mobilize leukemic cells in vivo, such that a plerixafor-nilotinib combination reduced the leukemia burden in mice significantly below the baseline level suppression exhibited by a moderate-to-high dose of nilotinib as single agent. These results support the idea of using CXCR4 inhibition in conjunction with targeted tyrosine kinase inhibition to override drug resistance in CML and suppress or eradicate residual disease.Leukemia advance online publication, 20 December 2011; doi:10.1038/leu.2011.360.

Dual antiplatelet therapy with aspirin and clopidogrel is among the most efficacious treatment for patients after acute coronary syndromes and for those who have had a percutaneous coronary intervention and coronary stent implantation. Patients who are treated with dual antiplatelet therapy are usually also ordered medications that reduce the secretion of gastric acid (such as H2 receptor blockers or proton pump inhibitors [PPIs]) in order to decrease the risk of gastrointestinal bleeding and dyspepsia. Numerous observational studies reported that omeprazole (a PPI) attenuates the antiplatelet activity and clinical effectiveness of clopidogrel and causes adverse cardiovascular events. Based on these findings, several medical agencies in the world have issued communications regarding the negative interaction between clopidogrel and PPIs, urging clinicians to evaluate the need for starting treatment with a PPI in patients taking clopidogrel. There are studies that reported contradicting findings, suggesting that there is no significant interaction between clopidogrel and PPIs. Only one prospective, randomized, double-blind, placebo-controlled clinical trial examined the interaction between clopidogrel and omeprazole and did not demonstrate cardiovascular harm among the patients who were treated with clopidogrel and omeprazole, as compared to those who were treated with clopidogrel and placebo. In this article, the authors review the current studies that reported a possible drug-drug interaction between clopidogrel and PPIs, particularly omeprazole.

Interactions between multiple myeloma (MM) cells and the BM microenvironment play a critical role in the pathogenesis of MM and in the development of drug resistance by MM cells. Selectins are involved in extravasation and homing of leukocytes to target organs. In the present study, we focused on adhesion dynamics that involve P-selectin glycoprotein ligand-1 (PSGL-1) on MM cells and its interaction with selectins in the BM microenvironment. We show that PSGL-1 is highly expressed on MM cells and regulates the adhesion and homing of MM cells to cells in the BM microenvironment in vitro and in vivo. This interaction involves both endothelial cells and BM stromal cells. Using loss-of-function studies and the small-molecule pan-selectin inhibitor GMI-1070, we show that PSGL-1 regulates the activation of integrins and downstream signaling. We also document that this interaction regulates MM-cell proliferation in coculture with BM microenvironmental cells and the development of drug resistance. Furthermore, inhibiting this interaction with GMI-1070 enhances the sensitization of MM cells to bortezomib in vitro and in vivo. These data highlight the critical contribution of PSGL-1 to the regulation of growth, dissemination, and drug resistance in MM in the context of the BM microenvironment.

Mammalian target of rapamycin (mTOR) is a downstream serine/threonine kinase of the PI3K/Akt pathway that integrates signals from the tumor microenvironment to regulate multiple cellular processes. Rapamycin and its analogs have not shown significant activity in multiple myeloma (MM), likely because of the lack of inhibition of TORC2. In the present study, we investigated the baseline activity of the PI3K/Akt/mTOR pathway TORC1/2 in MM cell lines with different genetic abnormalities. TORC1/2 knock-down led to significant inhibition of the proliferation of MM cells, even in the presence of BM stromal cells. We also tested INK128, a dual TORC1/2 inhibitor, as a new therapeutic agent against these MM cell lines. We showed that dual TORC1/2 inhibition is much more active than TORC1 inhibition alone (rapamycin), even in the presence of cytokines or stromal cells. In vitro and in vivo studies showed that p-4EBP1 and p-Akt inhibition could be predictive markers of TORC2 inhibition in MM cell lines. Dual TORC1/2 inhibition showed better inhibition of adhesion to BM microenvironmental cells and inhibition of homing in vivo. These studies form the basis for further clinical testing of TORC1/2 inhibitors in MM.

Waldenstrom macroglobulinemia (WM) is a low-grade B-cell lymphoma characterized by widespread involvement of the bone marrow with lymphoplasmacytic cells. In approximately 20% of patients, the malignant clone also involves the lymph nodes and induces hepatosplenomegaly. The mechanisms by which the tumor cells home to the bone marrow and preferentially reside in the marrow niches are not fully elucidated. In this review, we examine the role of the bone marrow microenvironment in the regulation of cell growth, survival and cell dissemination in WM. We also summarize specific regulators of niche-dependent tumor proliferation in WM. These include chemokines, adhesion molecules, Src/PI3K/Akt/mTOR signaling, NF-kB activation, and micro-RNA regulation in WM. Targeting these pathways in clinical trials could lead to significant responses in this rare disease.

PURPOSE: The Ephrin-receptors (Eph) are found in a wide range of cancers and correlate with metastasis. In this study, we characterized the role of Eph-B2 receptor in the interaction of Waldenstrom's Macroglobulinemia (WM) cells with the bone-marrow-microenvironment. EXPERIMENTAL DESIGN: We screened the activity of different RTKs in WM patients and found that Eph-B2 was over expression compared to control. Also, we tested the expression of Ephrin-B2 ligand on endothelial-cells and bone-marrow-stromal-cells isolate from WM patients. We then tested the role of Eph-B2/Ephrin-B2 interaction in the adhesion of WM cells to endothelial-cells and bone-marrow-stromal-cells; the cell-signaling induced by the co-culture in both the WM cells and the endothelial cells; WM cell proliferation, apoptosis and cell cycle in vitro and tumor progression in vivo; and in angiogenesis. RESULTS: Eph-B2-receptor was found to be activated in WM patients compared to control, with a 5-fold increase in CD19+ WM cells, and activated cell-adhesion signaling, including FAK, Src, p130, paxilin and cofilin, but decreased WM cell chemotaxis. Ephrin-B2 ligand was highly expressed on endothelial-cells and bone-marrow stromal-cells isolated from WM patients and on HUVEC cells, and induced signaling in the endothelial cells promoting adhesion and angiogenesis. Blocking of ephrin-B2 or Eph-B2 inhibited adhesion, cytoskeletal signaling, proliferation, and cell cycle in WM cells which was induced by co-culture with endothelial cells, and decreased WM tumor progression in vivo. CONCLUSION: Ephrin-B2/Eph-B2 axis regulates adhesion, proliferation, cell cycle and tumor progression in vivo through the interaction of WM with the cells in the BM microenvironment.

Lithium is the gold-standard treatment for bipolar disorder, a severe mental illness. A large body of evidence suggests that inflammation plays a role in the pathogenesis of bipolar disorder and that mood stabilizers exhibit anti-inflammatory properties. However, contradicting findings have also been reported. In this study, we examined the effects of lithium on LPS-induced inflammation in rat primary glia cells. Cells were pre-treated with lithium (1 or 10 mM) for 6 or 24 h, after which, inflammation was induced by the addition of LPS (for another 18 h) to the culture medium. Thereafter, medium was collected and cells were harvested for further analyses. Levels of TNF-α, IL1-β and PGE(2) were determined by ELISA and NO levels by the Griess reaction assay. Expression levels of cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS) were examined by Western blot analysis. We found that pre-treatment with lithium 10 mM (but not 1 mM) significantly reduced LPS-induced secretion of TNF-α, IL1-β, PGE(2) and NO. In addition, lithium significantly reduced the expression of COX-2 and iNOS. These findings indicate that lithium exhibits a potent anti-inflammatory effect. However, it's important to emphasize that this effect was obtained mainly under treatment with an extra-therapeutic concentration of the drug.

PURPOSE There is no standard of therapy for the treatment of Waldenström macroglobulinemia (WM), therefore there is a need for the development of new agents. Fibroblast growth factor receptor 3 (FGFR3) was shown to play a major role in several types in cancer. Dovitinib, an inhibitor of FGFR3, was effective in hematologic malignancies. In this study, we tested FGFR3 as a therapeutic target in WM and tested the effect of dovitinib on cell proliferation and apoptosis of WM cells in the context of BM microenvironment. Methods: The expression of FGFR3 in WM cells was tested using immunofluorescence and flow cytometry. Cell signaling in response to stimulation with FGF3 and stromal cells, and its inhibition by dovitinib was performed using immunoblotting. Cell survival and cell proliferation were assessed by MTT and BrdU assays. Apoptosis was measured by detection of APO-2.7 and cleavage of caspase-3 using flow cytometry. Cell cycle was performed by PI staining of cells and flow cytometry. The combinatory effect of dovitinib with other drugs was analyzed using Calcusyn software. The effect of dovitinib was tested in vivo. RESULTS FGFR3 was overexpressed in WM cells and its activation induced cell proliferation. Inhibition of FGFR3 with dovitinib decreased cell survival, increased apoptosis, and induced cell cycle arrest. Inhibition of FGFR3 by dovitinib reduced the interaction of WM to bone marrow components, and reversed its proliferative effect. Dovitinib had an additive effect with other drugs. Moreover, dovitinib reduced WM tumor progression in vivo. CONCLUSION We report that FGFR3 is a novel therapeutic target in WM, and suggest dovitinib for future clinical trial the treatment of patients with WM.