Prolonged exposure to a Mer ligand in leukemia: Gas6 favors expression of a partial Mer glycoform and reveals a novel role for Mer in the nucleus.

Mer tyrosine kinase is ectopically expressed in acute lymphoblastic leukemia and associated with enhanced chemoresistance and disease progression. While such effects are generally ascribed to increased engagement of oncogenic pathways downstream of Mer stimulation by its ligand, Gas6, Mer has not been characterized beyond the scope of its signaling activity. The present study explores Mer behavior following prolonged exposure to Gas6, a context similar to the Gas6-enriched microenvironment of the bone marrow, where a steady supply of ligand facilitates continuous engagement of Mer and likely sustains the presence of leukemic cells. Long-term Gas6 exposure induced production of a partially N-glycosylated form of Mer from newly synthesized stores of protein. Preferential expression of the partial Mer glycoform was associated with diminished levels of Mer on the cell surface and altered Mer localization within the nuclear-soluble and chromatin-bound fractions. The presence of Mer in the nucleus is a novel finding for this receptor, and the glycoform-specific preferences observed in each nuclear compartment suggest that glycosylation may influence Mer function within particular subcellular locales. Previous studies have established Mer as an attractive cancer biologic target, and understanding the complexity of its activity has important implications for potential strategies of Mer inhibition in leukemia therapy. Our results identify several novel features of Mer that expand the breadth of its functions and impact the development of therapeutic modalities designed to target Mer.

TAM receptors in leukemia: expression, signaling, and therapeutic implications.

In the past 30 years there has been remarkable progress in the treatment of leukemia and lymphoma. However, current treatments are largely ineffective against relapsed leukemia and, in the case of pediatric patients, are often associated with severe long-term toxicities. Thus, there continues to be a critical need for the development of effective biologically targeted therapies. The TAM family of receptor tyrosine kinases-Tyro3, Axl, and Mer-plays an important role in normal hematopoiesis, including natural killer cell maturation, macrophage function, and platelet activation and signaling. Furthermore, TAM receptor activation leads to upregulation of pro-survival and proliferation signaling pathways, and aberrant TAM receptor expression contributes to cancer development, including myeloid and lymphoid leukemia. This review summarizes the role of TAM receptors in leukemia. We outline TAM receptor expression patterns in different forms of leukemia, describe potential mechanisms leading to their overexpression, and delineate the signaling pathways downstream of receptor activation that have been implicated in leukemogenesis. Finally, we discuss the current research focused on inhibitors against these receptors in an effort to develop new therapeutic strategies for leukemia.

Comparison and usefulness of cardiac magnetic resonance versus computed tomography in infants six months of age or younger with aortic arch anomalies without deep sedation or anesthesia.

The present project investigated whether cardiac magnetic resonance (CMR) of aortic arch anomalies can be performed successfully in infants <6 months of age without the use of cardiac anesthesia or deep sedation. We performed a retrospective review of infants ≤6 months old from 2005 to 2009 who underwent either CMR or computed tomography angiography to investigate aortic arch abnormalities. The CMR procedure used a "feed and swaddle" protocol without deep sedation or cardiac anesthesia. Of the 52 infants referred for CMR, 24 underwent the feed and swaddle protocol (aged 2.6 ± 1.4 months). One patient awoke during the study, and examination of the remaining 23 yielded a definitive diagnosis (success rate 96%). The scanning time was 6.2 ± 3.1 minutes, with the large airways evaluation accounting for 1/2 the time. Single-shot axial steady-state free precession, in which the definitive diagnosis was made, accounted for 0.59 ± 0.3 minutes. Fifteen infants were diagnosed with a vascular ring. Of the 8 infants who underwent surgery, the diagnostic accuracy was 100%. During the same period, 19 patients, who had undergone computed tomography angiography (aged 1.67 ± 1.20 months), were referred for aortic arch evaluation. Of these 19 patients, 6 (32%) underwent sedation or anesthesia. The imaging time was 0.08 ± 0.06 minutes, significantly different from the CMR times (p <0.01). However, the overall room times (31.3 ± 22.3 and 35.8 ± 3.86 minutes, respectively) were not different between the CMR and angiographic groups. The radiation dose was 1.41 ± 1.03 mSv. In conclusion, CMR evaluation of aortic arch anomalies in children <6 months old can be successfully completed quickly using a feed and swaddle approach with high diagnostic accuracy. This protocol avoids the risks of sedation, as well as the radiation associated with computed tomography angiography.

In Silico Enhanced Restriction Enzyme Based Methylation Analysis of the Human Glioblastoma Genome Using Agilent 244K CpG Island Microarrays.

Genome wide methylation profiling of gliomas is likely to provide important clues to improving treatment outcomes. Restriction enzyme based approaches have been widely utilized for methylation profiling of cancer genomes and will continue to have importance in combination with higher density microarrays. With the availability of the human genome sequence and microarray probe sequences, these approaches can be readily characterized and optimized via in silico modeling. We adapted the previously described HpaII/MspI based Methylation Sensitive Restriction Enzyme (MSRE) assay for use with two-color Agilent 244K CpG island microarrays. In this assay, fragmented genomic DNA is digested in separate reactions with isoschizomeric HpaII (methylation-sensitive) and MspI (methylation-insensitive) restriction enzymes. Using in silico hybridization, we found that genomic fragmentation with BfaI was superior to MseI, providing a maximum effective coverage of 22,362 CpG islands in the human genome. In addition, we confirmed the presence of an internal control group of fragments lacking HpaII/MspI sites which enable separation of methylated and unmethylated fragments. We used this method on genomic DNA isolated from normal brain, U87MG cells, and a glioblastoma patient tumor sample and confirmed selected differentially methylated CpG islands using bisulfite sequencing. Along with additional validation points, we performed a receiver operating characteristics (ROC) analysis to determine the optimal threshold (p