The pediatric hydroxyurea phase III clinical trial (BABY HUG): challenges of study design.

Evidence of the laboratory benefits of hydroxyurea and its clinical efficacy in reducing acute vaso-occlusive events in adults and children with sickle cell anemia has accumulated for more than 15 years. A definitive clinical trial showing that hydroxyurea can also prevent organ damage might support widespread use of the drug at an early age. BABY HUG is a randomized, double-blind placebo-controlled trial to test whether treating young children ages 9-17 months at entry with a liquid preparation of hydroxyurea (20 mg/kg/day for 2 years) can decrease organ damage in the kidneys and spleen by at least 50%. Creation of BABY HUG entailed unique challenges and opportunities. Although protection of brain function might be considered a more compelling endpoint, preservation of spleen and renal function has clinical relevance, and significant treatment effects might be discernable within the mandated sample size of 200. Concerns about unanticipated severe toxicity and burdensome testing and monitoring requirements were addressed in part by an internal Feasibility and Safety Pilot Study, the successful completion of which was required prior to enrolling a larger number of children on the protocol. Concerns over recruitment of potentially vulnerable subjects were allayed by inclusion of a research subject advocate, or ombudsman. Finally, maintenance of blinding of research personnel was aided by inclusion of an unblinded primary endpoint person, charged with transmitting endpoint data and monitoring blood work locally for toxicity (ClinicalTrials.gov number, NCT00006400).

Severity of obstructive sleep apnea in children with sickle cell disease.

OBJECTIVE: To characterize polysomnographic (PSG) findings of children with sickle cell disease (SCD) suspected of having sleep disordered breathing (SDB). METHODS: Families of 100 consecutively referred children with SCD completed the Children's Sleep Habit Questionnaire during a routine visit to identify concerns regarding sleep habits and sleep behavior. Of these, 48 children were identified as displaying behaviors suspicious of SDB. Nineteen agreed to an overnight PSG. The results from the PSGs of the SCD with obstructive sleep apnea syndrome (OSAS) group (SCD-OSAS; group 1) were compared with the results of 10 age, sex, and ethnicity-matched patients identified as OSAS with no medical comorbidities (uncomplicated OSAS; group 2). RESULTS: SDB was identified in 79% of the SCD group. As compared with the uncomplicated OSAS group, the SCD with OSAS group displayed nocturnal desaturation with lower nadir values, of longer duration, with a 4-fold increased risk for oxygen desaturation below 85%, higher percentage of total sleep time with end-tidal carbon dioxide (ET CO2) values >50 mm Hg, with a 3.7-fold increased risk for spending more than 25% of total sleep time with ET CO2 more than 50 mm Hg and higher peak ET CO2 with a 7-fold increase for peak ET CO2 above 53 mm Hg. CONCLUSIONS: Children with SCD suspicious of SDB may have not only a higher incidence of OSAS, but also more severe nocturnal desaturation and hypercapnia as compared with children with uncomplicated OSAS.

Human CD34(+) and CD34(+)CD38(-) hematopoietic progenitors in sickle cell disease differ phenotypically and functionally from normal and suggest distinct subpopulations that generate F cells.

OBJECTIVE: Sickle cell disease (SCD) is remarkable for stress erythropoiesis. We investigated the progenitor populations contributing to erythroid stress. MATERIALS AND METHODS: We characterized hematopoietic progenitor cells in sickle bone marrow and sickle peripheral blood from patients with SCD compared to those in normal bone marrow. RESULTS: There were increased proportions of sickle bone marrow and sickle peripheral blood CD34(+) cells that coexpressed glycophorin A (GlyA), normally expressed late during erythroid differentiation when CD34 is down-regulated. Remarkably, increased numbers of CD34(+)CD38(-) hematopoietic progenitor cells from sickle bone marrow (p < 0.03) and sickle peripheral blood (p < 0.004) coexpressed GlyA, compared to normal bone marrow CD34(+)CD38(-) hematopoietic progenitor cells. At a molecular level, even the sickle bone marrow and sickle peripheral blood CD34(+)CD38(-) hematopoietic progenitor cells not expressing GlyA by fluorescence-activated cell sorting or reverse transcriptase-polymerase chain reaction expressed the erythroid-specific gene GATA-1, unlike normal bone marrow, suggesting desynchronized erythroid gene expression in the SCD hematopoietic progenitor cells. We also generated red blood cells in vitro from GlyA(+) and GlyA(-)CD34(+) cells. GlyA(+)CD34(+) produced more F cells (p < 0.02) and had lower clonogenicity (p < 0.01) and erythroid expansion potential. Increased F cells were generated only from sickle CD34(+) hematopoietic progenitor cells (p < 0.04), as occurs in vivo. CONCLUSION: Stress erythropoiesis in SCD has been postulated to accelerate erythropoiesis and production of F cells. Thus, CD34(+)CD38(-) expressing GlyA may represent the "stress progenitor" population. This is the first study characterizing CD34(+) and CD34(+)CD38(-) hematopoietic progenitor cells in sickle bone marrow, comparing them to sickle peripheral blood and normal bone marrow and using them to generate sickle red blood cells that recapitulate F cell production observed in vivo. We identified a unique population of GlyA(+)CD34(+) cells in SCD, which is in an accelerated erythroid differentiation pathway, has not down-regulated CD34 antigen expression, and predominantly generates F cells.

Terminal ileal atresia, total colonic aganglionosis, and thrombophilia.

Inherited thrombophilia, a predisposition for a hypercoagulable state, has been associated with cases of intestinal atresia. In this communication, we report a case of terminal ileal atresia and total colonic aganglionosis (Hirschsprung's disease), a rarely documented association, in a neonate who seemed to have a hypercoagulable state. The case stresses the need for recognition of this sequence of events in order to achieve optimal management.

Pediatric tumor cells express erythropoietin and a functional erythropoietin receptor that promotes angiogenesis and tumor cell survival.

Erythropoietin was traditionally considered an erythroid-restricted cytokine, but recent evidence indicates a broader role for it in nonhematopoietic tissues, specifically in neural development. Pediatric solid tumors are mostly developmental in origin, and more than 50% of the solid tumors are neural in origin. We found erythropoietin receptor and erythropoietin expression in common pediatric tumor cells: neuroblastomas, Ewing's sarcoma family of tumors, pediatric brain tumors (medulloblastoma, astrocytoma, and ependymoma), Wilms tumors, rhabdomyosarcomas, and hepatoblastomas (n = 24), and in cell lines derived from some of these tumors (n = 25). Expression of erythropoietin in tumor cell lines was hypoxia-inducible. Addition of exogenous erythropoietin to tumor cell lines expressing erythropoietin receptor increased nuclear DNA binding activity of nuclear factor kappa B and increased the expression of the antiapoptotic genes bcl-1, bcl-xL, and mcl-1. Additionally, exogenous erythropoietin increased production and secretion of angiogenic growth factors, vascular endothelial growth factor, or placenta growth factor from the tumor cell lines, which promoted endothelial cell proliferation and chemotaxis. Erythropoietin receptor expression that promotes tumor cell survival and releases angiogenic growth factors in pediatric tumors has not been previously described. Therefore, a careful evaluation of the impact of erythropoietin is warranted in vivo, in xenograft models of pediatric tumors, followed by evaluation in pediatric patients with cancer.

Placenta growth factor activates monocytes and correlates with sickle cell disease severity.

Sickle cell disease (SCD) results in chronic hypoxia and secondarily increased erythropoietin concentrations. Leukocytosis and activated monocytes are also observed in SCD in absence of infection or vaso-occlusion (steady state), the reasons for which are unknown. We found that erythroid cells produced placenta growth factor (PlGF), an angiogenic growth factor belonging to the vascular endothelial growth factor (VEGF) family, and its expression was induced in bone marrow CD34+ progenitor cells in the presence of erythropoietin. Furthermore, the steady state circulating PlGF levels in subjects with severe SCD (at least 3 vaso-occlusive crises [VOCs] per year) were 18.5 +/- 1.2 pg/mL (n = 9) compared with 15.5 +/- 1.2 pg/mL (n = 13) in those with mild SCD (fewer than 3 VOCs per year) and 11.3 +/- 0.7 pg/mL (n = 9) in healthy controls (P <.05), suggesting a correlation between PlGF levels and SCD severity. In addition, PlGF significantly increased mRNA levels of the proinflammatory cytochemokines interleukin-1beta, interleukin-8, monocyte chemoattractant protein-1, and VEGF in peripheral blood mononuclear cells (MNCs) of healthy subjects (n = 4; P <.05). Expression of these same cytochemokines was significantly increased in MNCs from subjects with SCD at steady state (n = 14), compared with healthy controls. Of the leukocyte subfractions, PlGF stimulated monocyte chemotaxis (P <.05, n = 3). Taken together, these data show for the first time that erythroid cells intrinsically release a factor that can directly activate monocytes to increase inflammation. The baseline inflammation seen in SCD has always been attributed to sequelae secondary to the sickling phenomenon. We show that PlGF contributes to the inflammation observed in SCD and increases the incidence of vaso-occlusive events.