Post-induction MRD by FCM and GATA1-PCR are significant prognostic factors for myeloid leukemia of Down syndrome.

Myeloid leukemia of Down syndrome (ML-DS) is associated with good response to chemotherapy, resulting in favorable outcomes. However, no universal prognostic factors have been identified to date. To clarify a subgroup with high risk of relapse, the role of minimal residual disease (MRD) was explored in the AML-D11 trial by the Japanese Pediatric Leukemia/Lymphoma Study Group. MRD was prospectively evaluated at after induction therapy and at the end of all chemotherapy, using flow cytometry (FCM-MRD) and GATA1-targeted deep sequencing (GATA1-MRD). A total of 78 patients were eligible and 76 patients were stratified to the standard risk (SR) group by morphology. In SR patients, FCM-MRD and GATA1-MRD after induction were positive in 5/65 and 7/59 patients, respectively. Three-year event-free survival (EFS) and overall survival (OS) rates were 93.3% and 95.0% in the FCM-MRD-negative population, and 60.0% and 80.0% in the positive population. Three-year EFS and OS rates were both 96.2% in the GATA1-MRD-negative population, and 57.1% and 71.4% in the positive population. Adjusted hazard ratios for associations of FCM-MRD or GATA1-MRD with EFS were 10.98 (p = 0.01) and 27.68 (p < 0.01), respectively. Detection of MRD by either FCM or GATA1 after initial induction therapy represents a significant prognostic factor for predicting ML-DS relapse

Genetic modification of primary natural killer cells overcomes inhibitory signals and induces specific killing of leukemic cells

Natural killer (NK) cells hold promise for improving the therapeutic potential of allogeneic hematopoietic transplantation, but their effectiveness is limited by inhibitory HLA types. We sought to overcome this intrinsic resistance by transducing CD56+CD3- NK cells with chimeric receptors directed against CD19, a molecule widely expressed by malignant B cells. An abundance of NK cells for transduction was secured by culturing peripheral blood mononuclear cells with K562 cells expressing the NK-stimulatory molecules 4-1BB ligand and interleukin 15, which yielded a median greater than 1000-fold expansion of CD56+CD3- cells at 3 weeks of culture, without T-lymphocyte expansion. Expression of anti-CD19 receptors linked to CD3ζ overcame NK resistance and markedly enhanced NK-cell-mediated killing of leukemic cells. This result was significantly improved by adding the 4-1BB costimulatory molecule to the chimeric anti-CD19-CD3ζ receptor; the cytotoxicity produced by NK cells expressing this construct uniformly exceeded that of NK cells whose signaling receptors lacked 4-1BB, even when natural cytotoxicity was apparent. Addition of 4-1BB was also associated with increased cell activation and production of interferon γ and granulocyte-macrophage colony-stimulating factor. Our findings indicate that enforced expression of signaling receptors by NK cells might circumvent inhibitory signals, providing a novel means to enhance the effectiveness of allogeneic stem cell transplantation

Reliability and validity of the Japanese version of the Paediatric Pain Profile for children with severe motor and intellectual disabilities.

Children with severe motor and intellectual disabilities experience chronic pain but cannot communicate verbally. However, no Japanese tool currently exists for assessing pain in this population. This study aimed to develop and evaluate the reliability and validity of a Japanese version of the Paediatric Pain Profile, which is a behavioral rating scale to assess pain in children with severe neurological disabilities. The sample comprised 30 children with severe motor and intellectual disabilities at three hospitals in Japan. Three specialist nurses rated low and high pain video scenes of the children (twice at 1-week intervals) using the Face, Legs, Activity, Cry, Consolability behavioral scale and a translated Japanese version of the Paediatric Pain Profile. On the basis of their ratings, we calculated the internal consistency, test-retest reliability, and intra- and inter-observer reliabilities of the Paediatric Pain Profile. Additionally, we assessed concurrent validity using the Face, Legs, Activity, Cry, Consolability behavioral scale and construct validity using low versus high pain scenes. Both internal consistency (low pain: alpha = 0.735; high pain: alpha = 0.928) and test-retest reliability (r = 0.846) of the Japanese version of the Paediatric Pain Profile were good. Intra-observer reliability was substantial (r = 0.748), whereas inter-observer reliability was only moderate (r = 0.529). However, the concurrent validity with Face, Legs, Activity, Cry, Consolability scores was good (r = 0.629) and construct validity was confirmed (p < 0.001). We confirmed the validity of the Japanese version of the Paediatric Pain Profile, but reliable pain assessment may require repeated ratings by the same person. To accurately assess pain in children with severe motor and intellectual disabilities, healthcare staff must be properly trained and become more skilled in using the Japanese version of the Paediatric Pain Profile

Neutrophils Induced Licensing of Natural Killer Cells

Natural killer (NK) cells acquire effector function through a licensing process and exert anti-leukemia/tumor effect. However, there is no means to promote a licensing effect of allogeneic NK cells other than cytomegalovirus reactivation-induced licensing in allogeneic hematopoietic stem cell transplantation in human. In mice, a licensing process is mediated by Ly49 receptors which recognize self-major histocompatibility complex class I. The distribution of four Ly49 receptors showed similar pattern in congenic mice, B10, B10.BR, and B10.D2, which have B10 background. Forty Gy-irradiated 2×106 B10.D2 cells including splenocytes, peripheral blood mononuclear cells in untreated mice, or granulocyte colony-stimulating factor treated mice were injected intraperitoneally into B10 mice. We found that murine NK cells were effectively licensed by intraperitoneal injection of donor neutrophils with its corresponding NK receptor ligand in B10 mice as a recipient and B10.D2 as a donor. Mechanistic studies revealed that NK cells showed the upregulation of intracellular interferon-γ and CD107a expression as markers of NK cell activation. Moreover, enriched neutrophils enhanced licensing effect of NK cells; meanwhile, licensing effect was diminished by depletion of neutrophils. Collectively, injection of neutrophils induced NK cell licensing (activation) via NK receptor ligand interaction

Mesenchymal stromal cell secretome up-regulates 47 kDa CXCR4 expression, and induce invasiveness in neuroblastoma cell lines.

Neuroblastoma accounts for 15% of childhood cancer deaths and presents with metastatic disease of the bone and the bone marrow at diagnosis in 70% of the cases. Previous studies have shown that the Mesenchymal Stromal Cell (MSC) secretome, triggers metastases in several cancer types such as breast and prostate cancer, but the specific role of the MSC factors in neuroblastoma metastasis is unclear. To better understand the effect of MSC secretome on chemokine receptors in neuroblastoma, and its role in metastasis, we studied a panel of 20 neuroblastoma cell lines, and compared their invasive potential towards MSC-conditioned-RPMI (mRPMI) and their cytokine receptor expression profiles. Western blot analysis revealed the expression of multiple CXCR4 isoforms in neuroblastoma cells. Among the five major isoforms, the expression of the 47 kDa isoform showed significant correlation with high invasiveness. Pretreatment with mRPMI up-regulated the expression of the 47 kDa CXCR4 isoform and also increased MMP-9 secretion, expression of integrin α3 and integrin β1, and the invasive potential of the cell; while blocking CXCR4 either with AMD 3100, a CXCR4 antagonist, or with an anti-47 kDa CXCR4 neutralizing antibody decreased the secretion of MMP-9, the expression of integrin α3 and integrin β1, and the invasive potential of the cell. Pretreatment with mRPMI also protected the 47 kDa CXCR4 isoform from ubiquitination and subsequent degradation. Our data suggest a modulatory role of the MSC secretome on the expression of the 47 kDa CXCR4 isoform and invasion potential of the neuroblastoma cells to the bone marrow