CD9 shapes glucocorticoid sensitivity in pediatric B-cell precursor acute lymphoblastic leukemia.

Resistance to glucocorticoids (GCs), the common agents for remission induction in pediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL), poses a significant therapeutic hurdle. Therefore, dissecting the mechanisms shaping GC resistance could lead to new treatment modalities. Here, we showed that CD9- BCP-ALL cells were preferentially resistant to prednisone and dexamethasone over other standard cytotoxic agents. Concordantly, we identified significantly more poor responders to the prednisone prephase among BCP-ALL patients with a CD9- phenotype, especially for those with adverse presenting features including older age, higher white cell count and BCR-ABL1. Furthermore, gain- and loss-of-function experiments dictated a definitive functional linkage between CD9 expression and GC susceptibility, as demonstrated by the reversal and acquisition of relative GC resistance in CD9low and CD9high BCP-ALL cells, respectively. Despite physical binding to the GC receptor NR3C1, CD9 did not alter its expression, phosphorylation or nuclear translocation but potentiated the induction of GC-responsive genes in GCresistant cells. Importantly, the MEK inhibitor trametinib exhibited higher synergy with GCs against CD9- than CD9+ lymphoblasts to reverse drug resistance in vitro and in vivo. Collectively, our results elucidate a previously unrecognized regulatory function of CD9 in GC sensitivity, and inform new strategies for management of children with resistant BCP-ALL.

Pharmacogenomic Profiling of Pediatric Acute Myeloid Leukemia to Identify Therapeutic Vulnerabilities and Inform Functional Precision Medicine.

Despite the expanding portfolio of targeted therapies for adults with acute myeloid leukemia (AML), direct implementation in children is challenging due to inherent differences in underlying genetics. Here we established the pharmacologic profile of pediatric AML by screening myeloblast sensitivity to approved and investigational agents, revealing candidates of immediate clinical relevance. Drug responses ex vivo correlated with patient characteristics, exhibited age-specific alterations, and concorded with activities in xenograft models. Integration with genomic data uncovered new gene-drug associations, suggesting actionable therapeutic vulnerabilities. Transcriptome profiling further identified gene-expression signatures associated with on- and off-target drug responses. We also demonstrated the feasibility of drug screening-guided treatment for children with high-risk AML, with two evaluable cases achieving remission. Collectively, this study offers a high-dimensional gene-drug clinical data set that could be leveraged to research the unique biology of pediatric AML and sets the stage for realizing functional precision medicine for the clinical management of the disease. SIGNIFICANCE: We conducted integrated drug and genomic profiling of patient biopsies to build the functional genomic landscape of pediatric AML. Age-specific differences in drug response and new gene-drug interactions were identified. The feasibility of functional precision medicine-guided management of children with high-risk AML was successfully demonstrated in two evaluable clinical cases. This article is highlighted in the In This Issue feature, p. 476.

R4 RGS proteins suppress engraftment of human hematopoietic stem/progenitor cells by modulating SDF-1/CXCR4 signaling.

Homing and engraftment of hematopoietic stem/progenitor cells (HSPCs) into the bone marrow (BM) microenvironment are tightly regulated by the chemokine stromal cell-derived factor-1 (SDF-1) and its G-protein-coupled receptor C-X-C motif chemokine receptor 4 (CXCR4), which on engagement with G-protein subunits, trigger downstream migratory signals. Regulators of G-protein signaling (RGS) are GTPase-accelerating protein of the Gα subunit and R4 subfamily members have been implicated in SDF-1-directed trafficking of mature hematopoietic cells, yet their expression and influence on HSPCs remain mostly unknown. Here, we demonstrated that human CD34+ cells expressed multiple R4 RGS genes, of which RGS1, RGS2, RGS13, and RGS16 were significantly upregulated by SDF-1 in a CXCR4-dependent fashion. Forced overexpression of RGS1, RGS13, or RGS16 in CD34+ cells not only inhibited SDF-1-directed migration, calcium mobilization, and phosphorylation of AKT, ERK, and STAT3 in vitro, but also markedly reduced BM engraftment in transplanted NOD/SCID mice. Genome-wide microarray analysis of RGS-overexpressing CD34+ cells detected downregulation of multiple effectors with established roles in stem cell trafficking/maintenance. Convincingly, gain-of-function of selected effectors or ex vivo priming with their ligands significantly enhanced HSPC engraftment. We also constructed an evidence-based network illustrating the overlapping mechanisms of RGS1, RGS13, and RGS16 downstream of SDF-1/CXCR4 and Gαi. This model shows that these RGS members mediate compromised kinase signaling and negative regulation of stem cell functions, complement activation, proteolysis, and cell migration. Collectively, this study uncovers an essential inhibitory role of specific R4 RGS proteins in stem cell engraftment, which could potentially be exploited to develop improved clinical HSPC transplantation protocols.

Double-unit unrelated cord blood transplantation for thalassemia major: Comparison with HLA-identical sibling bone marrow transplantation.

UCBT recipients with TM are at high risk of EF related to low number of stem cells and prior alloimmunization after multiple blood transfusions. Here, we evaluated the safety and efficacy of double-unit UCBT using TT-containing conditioning regimens in TM. Retrospective analysis of children who underwent double-unit UCBT for TM in the Prince of Wales Hospital between August 2007 and January 2017, and outcome of double-unit UCBT for TM was compared with outcome of HLA-matched sibling BMT. Ten patients, median age 4.2 years, received double-unit UCBT. All patients except one engrafted at a median of 19 days. None of the patients with successful engraftment had grade III or IV aGVHD. Among nine patients with successful engraftment, six of nine patients evaluable after day 100 developed cGVHD. All patients with cGVHD were well controlled after treatment with steroids and/or supportive care and maintained good quality of life. In comparison with patients receiving BMT, those given UCBT had slower platelet recovery, and more cGVHD. With a median follow-up of 272 months after BMT and 84 months after UCBT, the 8-year OS after BMT and UCBT was 92% and 90% (P = .84), whereas 8-year DFS after BMT and UCBT was 87% and 80% (P = .54). UCB could be an acceptable source of stem cells for transplantation of TM patients when HLA-matched family bone marrow donors are NA.

CD9 blockade suppresses disease progression of high-risk pediatric B-cell precursor acute lymphoblastic leukemia and enhances chemosensitivity.

CD9 has been implicated in cancer progression but its prognostic relevance and therapeutic potential in B-cell precursor acute lymphoblastic leukemia (BCP-ALL) are largely unknown. In a cohort of pediatric BCP-ALL patients, we found that CD9+ cases had a significantly lower 5-year relapse-free survival rate than CD9- cases. Multivariate analysis demonstrated that CD9 positivity independently predicted inferior survival outcomes, and could be applied with established prognostic features, including prednisone response and cytogenetic status, to refine patient stratification. Administration of CD9 antibody substantially suppressed disease progression in NOD/SCID mice xenografted with CD9+ cell lines and primary leukemic blasts from patients with high-risk and refractory BCP-ALL, without compromising hematopoietic stem cell engraftment. Combination of anti-CD9 with conventional chemotherapy further reduced leukemic burden and prolonged animal survival. Mechanistically, CD9 blockade inhibited leukemic cell proliferation, induced G0/G1 cell cycle arrest, activated p38, and enhanced chemotherapeutic agent-induced apoptosis. Further, CD9 physically interacted with integrin very late antigen-4, regulated affinity to vascular cell adhesion molecule-1, and was involved in leukemia-stroma interaction. Collectively, our study established CD9 as a new prognostic marker, validated the preclinical efficacy of CD9 antibody, and laid the foundation for clinical development of CD9-targeted therapy for high-risk and refractory pediatric BCP-ALL.

Indiscernible Benefit of Double-Unit Umbilical Cord Blood Transplantation in Children: A Single-Center Experience From Hong Kong.

Double-unit umbilical cord blood (DU-UCB) may extend the use of UCB transplantation and improve clinical outcomes. Data in the literature show that single-unit dominance happened in a vast majority of recipients, and the mechanism is unknown. We examined the clinical relevance and engraftment kinetics of DU-UCB transplant in 65 consecutive children who underwent unrelated single-unit (n = 25) and double-unit (n = 40) UCB transplantation for various hematological malignancies (n = 45) and nonmalignant disorders (n = 20). Our result showed no discernible benefit to children receiving double-unit transplant over those receiving single-unit transplant when the total nucleated cell (TNC) doses are ≥2.5 × 10(7)/kg, in terms of the hastening of the engraftment of neutrophils and platelets, reduction of nonengraftment, disease recurrence, early mortality, and graft-versus-host disease, despite significantly higher numbers of TNCs in double units. Further analyses demonstrated that the phenomena were not associated with underlying disease, duration of UCB storage, postthaw viability, HLA disparity, ABO incompatibility, gender, or doses of TNCs, CD34(+) cells, CD3(+) cells, or colony-forming units. Engrafting units in DU-UCB transplants were notably associated with higher CD34(+) cell dose. Chimerism studies demonstrated that single-unit dominance started before neutrophil engraftment in DU-UCB transplants. Data from the study suggested no advantage of infusing double-unit UCB, if an adequately dosed single-unit UCB is available. Successful prediction of the dominant graft would optimize algorithms of UCB selection and maximize the long-term engraftment of chosen units.

Isolated persistent elevation of alanine transaminase for early diagnosis of pre-symptomatic Wilson's disease in Chinese children.

BACKGROUND: Recent studies presented a contradictory approach for the investigation of pediatric patients with an isolated increase in alanine transaminase. While classical teaching advised for a thorough investigation, recent studies suggested the yield on further investigation was low and thus not necessary. Yet the approach to the same clinical problem may need to be different due to variable disease prevalence rates among different ethnic populations. For the population with a higher prevalence rate of genetic liver diseases like Wilson's disease, an abnormal liver function may be the first presenting feature for some patients. METHODS: We reviewed 10 Chinese children with Wilson's disease who were diagnosed at a presymptomatic stage because of an isolated persistent elevation of alanine transaminase. RESULTS: All 10 patients did not have overt symptoms of liver impairment or neurological deficit. They were picked up incidentally with an abnormal liver function test. All patients were started on treatment shortly after diagnosis, and they remained well and symptom-free on the latest follow-up. CONCLUSIONS: This case series illustrated that an isolated persistent elevation of alanine transaminase is an important clue to the early diagnosis of pre-symptomatic Wilson's disease. It is particularly relevant in the Asian population where the disease is more prevalent.

Humoral response to conjugate pneumococcal vaccine in paediatric oncology patients.

OBJECTIVE: Pneumococcal conjugate vaccine (PCV) is an effective way to prevent invasive pneumococcal diseases in high risk populations. The efficacy of this vaccine in paediatric oncology patients remains unknown. DESIGN AND SETTING: The authors evaluated the antibody response to seven pneumococcal serotypes in paediatric oncology patients given two doses of heptavalent PCV (PCV-7). RESULTS: Forty-four patients (20 males; 24 females) with median age 9.5 years were studied. After two doses of PCV-7, 86-100% of patients had protective antibody titres against the seven vaccine serotypes. Increases in geometric mean antibody concentrations ranged from 3.8-fold for serotype 19F to 85.8-fold for serotype 14. There was no documented invasive pneumococcal disease in our cohort during the study period. CONCLUSION: PCV can elicit protective antipneumococcal antibody responses in paediatric oncology patients.

The first reported HLCS gene mutation causing holocarboxylase synthetase deficiency in a Vietnamese patient.

BACKGROUND: Holocarboxylase synthetase deficiency is an inborn error of biotin metabolism leading to multiple carboxylase deficiency which is often biotin responsive. This disease is believed to be relatively common among the Asian population. METHODS: A 6-year-old Vietnamese boy presented with recurrent episodes of severe metabolic acidosis precipitated by intercurrent illnesses. An extensive skin rash was present since the onset of his illness. Multiple carboxylase deficiency was considered a likely diagnosis based on the history and the characteristic skin rash. RESULTS: This diagnosis was later confirmed by urine organic acid and molecular genetic studies. Urine organic acid showed characteristic excretion of glycine conjugates. Serum biotinidase activity was normal. Sequencing of the holocarboxylase synthetase gene revealed the patient being homozygous for a common mutation R508W. The patient showed a dramatic response to biotin within days of its administration. CONCLUSION: This case illustrates a potential highly treatable inborn error of metabolism that can be recognized on clinical grounds and its favorable response to biotin treatment.

Second malignant neoplasms in childhood cancer survivors in a tertiary paediatric oncology centre in Hong Kong, China.

BACKGROUND: Childhood cancer survivors were at risk of development of second malignant neoplasms. The aim of this study is to evaluate the incidence, risk factors and outcome of second malignant neoplasms in childhood cancer survivors in a tertiary paediatric oncology centre in Hong Kong, China. METHODS: We performed a retrospective review of patients with childhood cancer treated in Children's Cancer Centre in Prince of Wales Hospital, Hong Kong, China between May 1984 and June 2009. Case records of patients who developed second malignant neoplasms were reviewed. RESULTS: Totally 1374 new cases aged less than 21-year old were treated in our centre in this 25-year study period. Twelve cases developed second malignant neoplasms with 10-year and 20-year cumulative incidence of 1.3% (95% confidence interval 0.3% - 2.3%) and 2.9% (95% confidence interval 1.1% - 4.7%) respectively. Another 4 cases were referred to us from other centres for the management of second malignant neoplasms. In this cohort of 16 children with second malignant neoplasms, the most frequent second malignant neoplasms were acute leukemia or myelodysplastic syndrome (n = 6) and central nervous system tumor (n = 4). Median interval between diagnosis of primary and second malignant neoplasms was 7.4 years (range 2.1 - 13.3 years). Eight patients developed second solid tumor within the previous irradiated field. Radiotherapy significantly increased the risk of development of second solid tumor in patients with acute lymphoblastic leukemia (P = 0.027). Seven out of 16 patients who developed second malignant neoplasms had a family history of cancer among the first or second-degree relatives. Nine patients died of progression of second malignant neoplasms, mainly resulted from second central nervous system tumor and osteosarcoma. CONCLUSIONS: Cumulative incidence of second cancer in our centre was comparable to western countries. Radiotherapy was associated with second solid tumour among patients with acute lymphoblastic leukemia. Patients who developed second brain tumor and osteosarcoma had a poor outcome.

Recovery of humoral and cellular immunities to vaccine-preventable infectious diseases in pediatric oncology patients.

The recovery of antibodies to various vaccine-preventable infectious diseases, humoral and cellular immunity in pediatric oncology patients were evaluated by a prospective longitudinal study for 18 months. Lymphocyte subset (CD3+, CD4+, CD8+, CD16/56+, CD19+), CD4/CD8 ratio, immunoglobulin levels, antibodies to diphtheria, pertussis, tetanus, hepatitis B, measles, mumps, and rubella were measured serially at 6 months till 18 months after stopping all chemotherapy (including maintenance chemotherapy). Twenty-eight children (hematological malignancies, n = 14; solid tumors, n = 14) were studied. The median age was 7.0 +/- 3.8 years old (range 2.6-16.2 years old). Although there was significant increase in CD3+, CD4+, CD8+, CD19+ cells, IgG, IgA, and IgM levels (P < .05), CD4+ and CD8+ counts were still below the age-specific normal range at the end of study period. At 18 months after stopping chemotherapy, 11%, 15%, 60%, 30%, 49%, and 30% of subjects remained seronegative against diphtheria, tetanus, hepatitis B, measles, mumps, and rubella. This will evolve to a significant health care problem if no further intervention is implemented, as the survival rate of pediatric oncology patients improves significantly with the improvement in various cancer treatment protocols. Near complete immune recovery was demonstrated in the subjects. Significant proportion of subjects remained susceptible to vaccine-preventable infectious diseases up to 18 months after stopping all chemotherapy.

Oxidative challenge and glucose-6-phosphate dehydrogenase activity of preterm and term neonatal red blood cells.

BACKGROUND: Recent studies suggest that a low antioxidant level in preterm infants may predispose them to increased oxidative stress and results in hyperbilirubinemia, whereas glucose-6-phosphate dehydrogenase (G6PD) activity was found to be higher in preterm infants than in term infants. OBJECTIVES: To evaluate (1) the oxidative effect of alpha-naphthol on preterm and term red blood cells, and (2) the relationship between G6PD activity and the gestational age of these infants. METHODS: G6PD activities were determined in preterm and term infants by a standard diagnostic method. Whole blood samples were incubated with alpha-naphthol for 2 h and their pre- and post-challenged reduced glutathione (GSH) levels were quantified. RESULTS: The mean G6PD activity in preterm infants (n = 113; 13.52 +/- 0.19 U/g Hb; gestational age 30.67 +/- 0.28 weeks) was significantly higher than that in term infants (n = 100; 12.36 +/- 0.16 U/g Hb; gestational age 39.82 +/- 0.14 weeks; p < 0.001). A significantly negative correlation was demonstrated between gestational age and G6PD activity (r = -0.34, p < 0.001). GSH levels of preterm and term subjects were similar at baseline, but were significantly decreased upon challenge with alpha-naphthol (p < 0.001). The percentage reduction in GSH levels was similar in the various gestational age groups. CONCLUSIONS: Our data show that G6PD activities had a negative correlation with gestational age of G6PD-normal infants. The similar response of preterm and term erythrocytes to an alpha-naphthol challenge indicates the manifestation of an active anti-oxidative pathway mediated by cellular GSH.

Humoral immune response after post-chemotherapy booster diphtheria-tetanus-pertussis vaccine in pediatric oncology patients.

BACKGROUND: The role of post-chemotherapy booster vaccination in pediatric oncology children remains to be established. In this randomized controlled study, we studied the effect of immune responses to diphtheria-tetanus-pertussis (DTP) booster vaccination in children 6 months after completing chemotherapy. METHODS: Children 1-18 years old with chemotherapy completed for 6 months (baseline) were eligible. Subjects were randomized into vaccine and control group. In the former, three doses of DTP vaccine (Aventis Pasteur Inc., Lyon, France) were administered. IgG antibody titers against diphtheria, tetanus, pertussis, hepatitis B, measles, mumps, and rubella antibodies were measured serially in vaccine and control groups. Subsets of circulating lymphocytes (CD3(+), CD4(+), CD8(+), CD19(+), and CD16/56(+)) were quantified by flow cytometry using fluorescence-labeled monoclonal antibodies. RESULTS: Fifty-six children (28 vaccinees; 28 controls) were enrolled. Protective antibody levels against diphtheria, tetanus, pertussis were found at baseline in 83.6%, 96.5%, 96.1% of them respectively. After three doses of DTP, all vaccinees demonstrated a sustain rise in antibody levels and the antibody titers were significantly higher than control group. 35.8% of subjects were susceptible to measles mumps and rubella infection and 69% showed anti-HBs antibody titer less than protective level up to 18 months after stopping chemotherapy. CONCLUSIONS: Post-chemotherapy booster vaccinations produced a strong and sustained effect in humoral immunity against vaccine-preventable infectious diseases.

Haemophagocytic lymphohistiocytosis in Hong Kong children.

Haemophagocytic lymphohistiocytosis is a rare but potentially fatal disease. Diagnosing this disease may be difficult and is often delayed because the clinical presentation mimics other conditions like severe sepsis, hepatic failure and malignancies. We reviewed the clinical presentations, response to treatment, and outcomes of children diagnosed with haemophagocytic lymphohistiocytosis from 1991 to 2006 in a Hong Kong tertiary paediatric haematology centre. All patients had typical presentations with prolonged fever, organomegaly, and pancytopaenia. Six children had hepatic dysfunction and two had neurological symptoms. The median time from disease onset to diagnosis was 21 days. Elevated serum ferritin levels and evidence of haemophagocytosis in bone marrow examinations aided diagnosis. The overall mortality was 57%. Three patients who presented in the first few years studied had relatively long lag times between disease onset and definitive treatment; all died of active disease. Three patients diagnosed more recently were given timely treatment using the haemophagocytic lymphohistiocytosis-94 protocol of etoposide and dexamethasone, with or without cyclosporin. All three achieved remission, but two had a recurrence and one died during the recurrence.

The plant mannose-binding lectin NTL preserves cord blood haematopoietic stem/progenitor cells in long-term culture and enhances their ex vivo expansion.

Ex vivo expansion of haematopoietic stem and progenitor cells in cytokine combinations is effective in promoting differentiation and proliferation of multilineage progenitor cells, but often results in reduction of self-renewable stem cells. This study investigated the effect of a mannose-binding lectin, NTL, purified from Narcissus tazetta var. chinensis, on prolonged maintenance and expansion of cord blood CD34+ cells. Our results showed that the presence of NTL or Flt-3 ligand (FL) significantly preserved a population of early stem/progenitor cells in a serum- and cytokine-free culture for 35 d. The effect of NTL on the ex vivo expansion of CD34+ cells in the presence of stem cell factor, thrombopoietin (TPO) and FL was also investigated. NTL-enhanced expansion of early progenitors (CD34+, CD34+CD38-, mixed colony-forming units and CFU-GEMM) and committed progenitor cells (granulocyte CFU, erythroid burst-forming units/CFU and megakayocyte CFU) after 8 and 12 d of culture. Six weeks after transplanting 12 d-expanded cells to non-obese diabetic severe combined immunodeficient mice, increased engraftment of human CD45+ cells was observed in the bone marrow of animals that received NTL-treated cells. The dual functions of NTL on long-term preservation and expansion of early stem/multilineage progenitor cells could be developed for applications in various cell therapy strategies, such as the clinical expansion of CD34+ cells for transplantation.

Erratum to: Mutational analysis of 65 Wilson disease patients in Hong Kong Chinese: Identification of 17 novel mutations and its genetic heterogeneity.

The correct name of the eighth author should be given as Sik-To Lai, not Jak-Yiu Lai.

Mutational analysis of 65 Wilson disease patients in Hong Kong Chinese: identification of 17 novel mutations and its genetic heterogeneity.

Wilson disease (WD), an autosomal recessive disorder of copper transport, is the most common inherited liver disorder in Hong Kong Chinese. This was the first local study to elucidate the molecular basis and establish an effective DNA-based diagnostic protocol. The ATP7B genes of 65 patients were amplified by polymerase chain reaction (PCR) and sequenced. Haplotype analysis was performed using D13S301, D13S314, and D13S316. The p.L770L/p.R778L status in 660 subjects was determined to estimate WD prevalence. Allele age of p.R778L was determined by the smallest homozygosity region between D13S301 and D13S270. We identified 42 different mutations with 17 being novel. p.R778L (17.3%) was the most prevalent. Exons 2, 8, 12, 13, and 16 harbored 70% mutations. Thirty-two haplotypes were associated with WD chromosomes. The estimated prevalence rate was 1 in 5,400. Three out of 660 normal subjects had p.L770L/p.R778L. In the remaining 657 individuals, neither p.L770L nor p.R778L was found. We characterized a Hong Kong Chinese-specific ATP7B mutation spectrum with great genetic diversity. Exons 2, 8, 12, 13, and 16 should be screened first. The perfect linkage disequilibrium suggested that p.R778L and its private polymorphism p.L770L originated from a single ancestor. This East-Asian-specific mutation p.R778L/p.L770L is aged at least 5,500 years.

Promoting effects of serotonin on hematopoiesis: ex vivo expansion of cord blood CD34+ stem/progenitor cells, proliferation of bone marrow stromal cells, and antiapoptosis.

Serotonin is a monoamine neurotransmitter that has multiple extraneuronal functions. We previously reported that serotonin exerted mitogenic stimulation on megakaryocytopoiesis mediated by 5-hydroxytryptamine (5-HT)2 receptors. In this study, we investigated effects of serotonin on ex vivo expansion of human cord blood CD34+ cells, bone marrow (BM) stromal cell colony-forming unit-fibroblast (CFU-F) formation, and antiapoptosis of megakaryoblastic M-07e cells. Our results showed that serotonin at 200 nM significantly enhanced the expansion of CD34+ cells to early stem/progenitors (CD34+ cells, colony-forming unit-mixed [CFU-GEMM]) and multilineage committed progenitors (burst-forming unit/colony-forming unit-erythroid [BFU/CFU-E], colony-forming unit-granulocyte macrophage, colony-forming unit-megakaryocyte, CD61+ CD41+ cells). Serotonin also increased nonobese diabetic/severe combined immunodeficient repopulating cells in the expansion culture in terms of human CD45+, CD33+, CD14+ cells, BFU/CFU-E, and CFU-GEMM engraftment in BM of animals 6 weeks post-transplantation. Serotonin alone or in addition to fibroblast growth factor, platelet-derived growth factor, or vascular endothelial growth factor stimulated BM CFU-F formation. In M-07e cells, serotonin exerted antiapoptotic effects (annexin V, caspase-3, and propidium iodide staining) and reduced mitochondria membrane potential damage. The addition of ketanserin, a competitive antagonist of 5-HT2 receptor, nullified the antiapoptotic effects of serotonin. Our data suggest the involvement of serotonin in promoting hematopoietic stem cells and the BM microenvironment. Serotonin could be developed for clinical ex vivo expansion of hematopoietic stem cells for transplantation. Disclosure of potential conflicts of interest is found at the end of this article.

Small peptide analogue of SDF-1alpha supports survival of cord blood CD34+ cells in synergy with other cytokines and enhances their ex vivo expansion and engraftment into nonobese diabetic/severe combined immunodeficient mice.

The SDF-1/CXCR4 axis has been implicated in the chemotaxis, homing, mobilization, and expansion of hematopoietic stem and progenitor cells. We studied the effects of a SDF-1 peptide analogue CTCE-0214 on the survival of cord blood CD34+ cells in culture, expansion, and engraftment of expanded cells in the nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse model. Our results demonstrated that CTCE-0214 synergized with thrombopoietin (TPO), stem cell factor (SCF), or flt-3 ligand (FL) on the survival of stem and progenitor cells in culture. Adding CTCE-0214 at a low concentration (0.01 ng/ml) for 4 days together with TPO, SCF, and FL significantly enhanced ex vivo expansion of CD34+ cells to subsets of primitive (CD34+CD38- cells, colony-forming unit-mixed [CFU-GEMMs]), erythroid (CFU-Es), myeloid (CFU-GMs), and megakaryocytic (CD61+CD41+ cells, CFU-MKs) progenitors, as well as their multilineage engraftment in NOD/SCID mice. Interestingly, the short exposure of expanded cells to CTCE-0214 (100 and 500 ng/ml) for 4 hours did not increase the quantity of progenitor cells but enhanced their engraftment capacity. The proportion of CD34+ cells expressing surface CXCR4 was decreased, but the overall number of this population increased upon expansion. The small peptide analogue of SDF-1 could be developed for ex vivo expansion and improving engraftment of cord blood transplantation.

Arsenic trioxide mediates intrinsic and extrinsic pathways of apoptosis and cell cycle arrest in acute megakaryocytic leukemia.

Arsenic trioxide (ATO) induces apoptosis in a range of solid tumors and leukemia cells, and has been clinically applied for the treatment of acute promyelocytic leukemia with confirmed efficacy. Acute megakaryocytic leukemia (AMKL) is an aggressive malignancy with poor prognosis, if bone marrow transplantation is not possible. In this study, we applied flow cytometry, Western blot analysis and microarray techniques to investigate the effects of ATO on apoptosis and the cell division cycle of AMKL cell lines CHRF-288-11 and MEG-01. Our data demonstrated that ATO is a potent agent against AMKL as indicated by apoptotic markers, Annexin V and caspase-3. ATO activated the intrinsic (mitochondrial) pathway of apoptosis, which involved disrupting mitochondrial membrane potential, increased Bax/Bcl-2 ratio and caspase-9 activation, as well as the extrinsic (death receptor) pathway mediated by Fas and caspase-8 activation. We provided the first evidence that ATO stimulated expressions of CD137 mRNA and protein, which might be relevant to the extrinsic mechanism. ATO induced delays of cell cycle progression at S phase and arrest at G2/M phase of AMKL cells, but caspase-3 expression appeared not to be phase-specific. The multiple-signaling mechanism of ATO warrants it a potential agent to incorporate in the treatment regimen of AMKL.