[Study on Down-regulation of Interleukin-1ß Secretion by Inhibiting ABCC1/MRP1 Transporter].

OBJECTIVE: To screen interleukin (IL)-1ß secretion-related membrane transporters by macrophage experiment in vitro and conventional knockout mice. METHODS: THP-1 cell line was differentiated to obtain human THP-1-derived macrophages, and the primary macrophages were obtained from human peripheral blood. FVB wild-type mice with the same sex and age were used as the controls of MRP1 knockout mice. The macrophages in abdominal cavity and bone marrow of mice were cultivated. The cells were treated with ABCC1/MRP1, ABCG2/BCRP, ABCB1/P-gp, OATP1B1, and MATE transporter inhibitors, then stimulated by lipopolysaccharide and adenosine triphosphate. The secretion level of IL-1ß was detected by ELISA, Western blot, and immunofluorescence. RESULTS: After inhibiting ABCC1/MRP1 transporter, the secretion of IL-1ß decreased significantly, while inhibition of the other 4 transporters had no effect. In animal experiment, the level of IL-1ß secreted by macrophages in bone marrow of MRP1 knockout mice was significantly lower than control group (P < 0.05). CONCLUSION: ABCC1/MRP1 transporter is a newly discovered IL-1ß secretion pathway, which is expected to become a new target for solving clinical problems such as cytokine release syndrome.

The predictive utility of cytokines, procalcitonin and C-reactive protein among febrile pediatric hematology and oncology patients with severe sepsis or septic shock.

Severe sepsis and septic shock are life-threatening for pediatric hematology and oncology patient receiving chemotherapy. Th1/Th2 cytokines, C-reactive protein (CRP), and procalcitonin (PCT) are all thought to be associated with disease severity. The aim of this study was to prospectively verify the utility of Th1/Th2 cytokines and compare them with PCT and CRP in the prediction of adverse outcomes. Data on patients were collected from January 1, 2011, to December 31, 2020. Blood samples were taken for Th1/Th2 cytokine, CRP, and PCT measurements at the initial onset of infection. Severe infection (SI) was defined as severe sepsis or septic shock. Th1/Th2 cytokine levels were determined by using flow cytometric bead array technology. In total, 7,735 febrile episodes were included in this study. For SI prediction, the AUCs of IL-6, IL-10 and TNF-α were 0.814, 0.805 and 0.624, respectively, while IL-6 and IL-10 had high sensitivity and specificity. IL-6 > 220.85 pg/ml and IL-10 > 29.95 pg/ml had high odds ratio (OR) values of approximately 3.5 in the logistic regression. Within the subgroup analysis, for bloodstream infection (BSI) prediction, the AUCs of IL-10 and TNF-α were 0.757 and 0.694, respectively. For multiorgan dysfunction syndrome (MODS) prediction, the AUC of CRP was 0.606. The AUC of PCT for mortality prediction was 0.620. In conclusion, IL-6 and IL-10 provide good predictive value for the diagnosis of SI. For children with SI, IL-10 and TNF-α are associated with BSI, while CRP and PCT are associated with MODS and death, respectively.

Prognostic performance of IL-6 and IL-10 in febrile pediatric hematology/oncology patients with normal procalcitonin.

INTRODUCTION: It is important to predict adverse outcomes in febrile children with hematology/oncology diseases. Procalcitonin (PCT) is a promising biomarker for the prediction of infection severity, but further studies have revealed its performance in excluding adverse outcomes of infection. IL-6 and IL-10 were reported to have a close association with those infection outcomes. The aim of the study was to investigate the performance of IL-6 and IL-10 in febrile pediatric hematology/oncology patients with normal PCT. METHODS: This was a retrospective study conducted in a tertiary children's hospital in China over the past ten years. Inflammatory biomarkers, including IL-6, IL-10, PCT and C-reactive protein (CRP), were detected at the onset of infection. Separate analyses were conducted in patients with neutropenia and without neutropenia. RESULTS: In total, 5987 febrile cases were enrolled. For patients with neutropenia, IL-6, IL-10 and PCT were significantly increased in patients with bloodstream infection (BSI), gram-negative bacteremia (GNB) and severe sepsis (SS), but only IL-6 and IL-10 were predictive of GNB and SS. For patients without neutropenia, IL-6, IL-10 and PCT were significantly increased in patients with BSI, GNB and SS, but no biomarkers were predictive of adverse outcomes. All biomarkers failed to exclude patients with fever of unknown origin or upper respiratory infection/bronchitis in patients with neutropenia. CONCLUSIONS: IL-6 and IL-10 could be predictors for GNB and SS in febrile patients with neutropenia and had some association with unfavorable outcomes in febrile patients without neutropenia. All biomarkers failed to exclude patients with fever of unknown origin or upper respiratory infection/bronchitis.

The effect of the plasma methotrexate concentration during high-dose methotrexate therapy in childhood acute lymphoblastic leukemia.

Two hundred and thirty-one acute lymphoblastic leukemia (ALL) children with 1376 high-dose methotrexate (HD-MTX) courses (3-5 g/m2) were enrolled to analyze the influence of the plasma MTX concentration (CMTX) in ALL. The 24-h target peak CMTX (C24h) was set at 33 µmol/l for low-risk (LR) and 65 µmol/l for intermediate/high-risk (IR/HR) groups. The median C24h was 42.0 µmol/l and 69.7 µmol/l for LR and IR/HR groups, respectively. MTX excretion delay was observed in 14.6% of courses, which was more frequent in IR/HR groups (56.9% vs. LR group 40.2%, p = .014) and T-ALL patients (82.6% vs. B-ALL 47.1%, p = .001). MTX-related toxicities were more common in courses with MTX excretion delay. However, survival between the patients who failed to reach the target C24h or not, with or without MTX excretion delay, was comparable. These findings suggest that, owing to the effectiveness of risk stratification chemotherapy, CMTX does not exert an independent influence on the prognosis of childhood ALL.

[Role of Flow Cytometry in the Diagnosis of Chronic Myeloid Leukemia].

OBJECTIVE: To analyze the immunological phenotype of chronic myeloid leukemia (CML), and explore its characteristics and significance. METHODS: The immunophenotypes of 40 CML children and 40 controls were analyzed by multicolor flow cytometry. CD45/SSC, as the basic gate, was used to delineate neutrophils. Then, the distribution of cluster differentiation (CD) molecules on the surface of granulocytes was analyzed in three ranges (≥1%, ≥5%, and ≥20%), and the expression rates of CD molecules (≥1% included in the statistical analysis) and the mean fluorescence intensity (MFI) were compared between the two groups. RESULTS: The proportion of granulocytes in the CML group was (82.1±6.4)%, which was significantly higher than (57.8±11.8)% in the control group (P <0.001). The expression rates of CD15/CD11b/CD33/CD13 in CML and control groups were high, and both distributed in the range of ≥20%. The differentiation trajectory of CD33/CD13 was normal and there were no significant differences in the expression rate and MFI between the two groups. However, both the expression rate of CD11b and CD15 MFI in the CML group were significantly lower than those in the control group (P <0.001). There were no significant differences in the expression rate and MFI of CD10 between the two groups, and the expression levels of CD10 between the two groups were consistent in different distributions. In the CML group, there was a large number of cases with abnormal high expression of CD56, 52.5% of the cases had a CD56 expression rate of ≥5%, and 42.5% had a CD56 expression rate of ≥20%, while the control group did not express CD56 (<1%). The expression distribution of CD117 was different between the two groups. In the range of expression rate ≥5%, there were 35.0% cases in the CML group, while only 2.5% in the control group. The expression rate of CD117 in the CML group was higher than that in the control group (P <0.001), though there was no significant difference in MFI. CONCLUSION: The immunophenotyping of CML is characterized by increased proportion of mature neutrophils, decreased CD15 MFI, decreased proportion of CD11b and abnormal high expression of CD56 and CD117. Flow cytometric analysis of immunophenotype can effectively distinguish normal granulocytes from chronic granulocytes, and help in the diagnosis of CML.

[Effect of Human Bone Marrow Mesenchymal Stem Cells with Ectopic High OCT4 Expression on T Lymphocyte Function].

OBJECTIVE: To explore the effect of human bone marrow mesenchymal stem cells (MSCs) with ectopic high OCT4 expression on T-cell proliferation, activation and secretion in vitro. METHODS: Peripheral blood mononuclear cells were isolated from healthy children. Anti-CD3 and anti-CD28 monoclonal antibodies were used to activate T lymphocytes, which were stimulated by interleukin (IL)-2 for one week in vitro. Then MSCs with ectopic high OCT4 expression (MSC-OCT4) were co-cultured with activated T lymphocytes. After one week of co-culture, the supernatant was collected and the levels of Th1/Th2 cytokines [IL-2, IL-4, IL-6, IL-10, tumor necrosis factor (TNF)-α and interferon (IFN)-γ] were determined by flow cytometry. The lymphocytes after one week of co-culture were collected and counted by Countstar software. After the proportions of activated/inactivated T cell subsets were determined by flow cytometry, the absolute lymphocyte counts were calculated and expressed as mean ± standard deviation. RESULTS: Compared with control T cell alone culture group, the proliferation of CD3+ T cells, CD3+CD4+ T cells, and CD3+CD8+ T cells were significantly inhibited in MSC group and MSC-OCT4 group. Compared with MSC, MSC-OCT4 could inhibit CD3+CD8+ T cell proliferation better (P =0.049), and mainly inhibited early T cell activation. Compared with control T cell alone culture group, the levels of IL-2 and INF-γ were significantly down-regulated both in MSC group and MSC-OCT4 group.After co-culture with T cells for one week, the level of IL-6 significantly increased in MSC group and MSC-OCT4 group compared with that before co-culture. Compared with control MSC group, MSC-OCT4 group had higher viable cell numbers after 1 week of co-culture (P =0.019), and could resist the inhibition of proliferation by higher concentration of mitomycin C. CONCLUSION: Both MSC and MSC-OCT4 can inhibit the proliferation and activation of IL-2-stimulated T cells in vitro. After overexpression of OCT4, MSC has better proliferation ability in vitro and can inhibit the proliferation of CD3+CD8+ T cells more effectively, which may have a better and more lasting immunosuppressive ability to regulate the balance of Th1/Th2.

Clinical features and prognostic implications of ecotropic viral integration site 1 (EVI1) in childhood acute lymphoblastic leukemia.

In contrast to the extensive knowledge on EVI1 in myeloid malignancies, few data are available on the EVI1 transcript in pediatric ALL. The purpose of this study was to examine the clinical and biological significance of EVI1 and validate its prognostic significance in pediatric patients with ALL. Here, we examined the clinical and biological significance of EVI1 expression, as measured by real-time polymerase chain reaction (PCR) in 837 children with newly diagnosed ALL treated on the National Protocol of Childhood Leukemia in China (NPCLC)-ALL-2008 protocol, and aimed to explore their prognostic significance in pediatric ALL patients. The EVI1 expression was detected in 27 of 837 (3.2%) patients. No statistically significant differences in prednisone response, complete remission (CR) rates and relapse rates were found between EVI1 overexpression (EVI1+) group and EVI1- group. Moreover, we found no significant difference in event-free survival (EFS) and overall survival (OS) between these two groups, also multivariate analysis did not identify EVI1+ as an independent prognostic factor. In the subgroup analysis, there was no difference in clinical outcome between EVI1+ and EVI1- patients in standard­risk (SR), intermediate-risk (IR) and high-risk (HR) groups. In the minimal residual disease (MRD)<10-4 group, EVI1+ patients have significantly lower EFS and OS rates compared to EVI1- patients. Further large­scale and well­designed prospective studies are required to confirm the results in the future.

[Advances in Structural Designs of Chimeric Antigen Receptor T Cells--Review].

Although chimeric antigen receptor (CAR)-T therapy has produced remarkable clinical responses for patients with relapsed or refractory hematological malignancies, setbacks were experienced, including antigen escape and heterogeneity, its efficacy and safety issues. In recent years, researchers at home and abroad are addressing the current obstacles by digging deeply into structural optimization of CAR gene in order to solve the problems of CAR-T cell therapy. In this review, we mainly illustrate the ectodomain structure, transmemberane domain, and endodomain structure, and new designs which promote persistence of CAR-T cells in vivo, so as to provide new ideas for improving the safety and the efficacy of CAR-T cell therapy.

Metagenomic next-generation sequencing for the diagnosis of fever of unknown origin in pediatric patients with hematological malignancy.

BACKGROUND AND AIMS: The aim of this study was to evaluate the performance of metagenomic next-generation sequencing (mNGS) in identifying microbiological etiologies in pediatric patients with hematological malignancies undergoing fever of unknown origin (FUO). METHODS: A total of 147 children with hematological malignancy suffering febrile diseases without definite microbiological etiologies under conventional tests were enrolled. The clinical record, serum inflammatory biomarkers and mNGS results were analyzed. RESULTS: At least one microorganism was identified by mNGS in 112 of 147 patients (76.2 %). Two or more types of organisms were detected simultaneously in 35.7 % (40/112) of samples. Of the 112 cases with positive mNGS results, the reported microorganisms were considered as etiologies of fever in 50 (44.6 %) cases. The initial antimicrobial regimens were adjusted according to the mNGS results in 48 cases, with 41 patients' febrile diseases resolved. Totally, 27.9 % (41/147) of patients benefit from mNGS. High IL-6 (>390 pg/mL) level was associated with bacterial infection and could help to interpret the results of mNGS. CONCLUSION: mNGS is a novel approach to determine the microbiological etiology of FUO in hematological malignancy patients, which benefits about a quarter of all patients tested. Integration of IL-6 can improve the diagnostic precision of bacterial infection.

Integration of Interleukin-6 Improves the Diagnostic Precision of Metagenomic Next-Generation Sequencing for Infection in Immunocompromised Children.

The performance of metagenomic next-generation sequencing (mNGS) in identifying pathogens in immunocompromised children was not very clear. The purpose of this study is to assess the performance of mNGS in this population and to investigate whether the integration of serum cytokines and mNGS assay could improve diagnostic accuracy. We retrospectively collected the clinical data of pediatric patients who suffered febrile diseases and underwent mNGS determination simultaneously in the department of hematology/oncology between January 2019 and March 2021. Specimens were sent for conventional microbiological test (CMT), mNGS, and serum cytokine measurement in parallel. A total of 258 episodes of febrile diseases were enrolled, mNGS was positive in 224 cases, while CMT was positive in 78 cases. mNGS and CMT were both positive in 70 (27.1%) cases and were both negative in 26 (10.1%) cases. There were 154 (59.7%) cases positive by mNGS only while 8 (3.1%) were positive by CMT only. It was common that two or more pathogens were simultaneously detected by mNGS in a single specimen, with only 61 tests identified a single organism. Whether the organisms reported by mNGS were the microbiological etiology of infection was evaluated. Of the 224 cases with positive mNGS results, 135 (58.4%), 30 (13.0%), and 59 (28.6%) were considered as "probable," "possible," and "unlikely," respectively. Patients with high IL-6 (≥ 390 pg/ml) were likely to be bacterial infection. Although mNGS reported mixed pathogens, 84.6% (33/39) and 83.3% (10/12) of patients presenting high IL-6 were confirmed as bacterial infection in the training and validation cohort, respectively. In conclusion, mNGS analysis demonstrates promising diagnostic potential in rapidly identifying clinically relevant pathogens. Given the detection of many clinically irrelevant organisms, the integration of IL-6 improves the precision of mNGS results interpretation.

Simple Evaluation of Clinical Situation and Subtypes of Pediatric Hemophagocytic Lymphohistiocytosis by Cytokine Patterns.

Background: Hemophagocytic lymphohistiocytosis (HLH) is a rapidly fatal disease caused by immune dysregulation. Early initiation of treatment is imperative for saving lives. However, a laboratory approach that could be used to quickly evaluate the HLH subtype and clinical situation is lacking. Our previous studies indicated that cytokines such as interferon (IFN)-γ and interleukin (IL)-10 were helpful for the early diagnosis of HLH and were associated with disease severity. The purpose of this study is to clarify the different cytokine patterns of various subtypes of pediatric HLH and to investigate the role of cytokines in a simple evaluation of disease feature. Patients and Methods: We enrolled 256 pediatric patients with newly diagnosed HLH. The clinical features and laboratory findings were collected and compared among different subtypes of HLH. A model integrating cytokines was established to stratify HLH patients into different clinical groups. Results: Twenty-seven patients were diagnosed with primary HLH (pHLH), 179 with EBV-HLH, and 50 with other causes. The IL-6, IL-10, and IFN-γ levels and the ratios of IL-10 to IFN-γ were different among EBV-HLH, other infection-associated HLH, malignancy-associated HLH, familial HLH, and X-linked lymphoproliferative disease. Patients with the ratio of IL-10 to IFN-γ >1.33 and the concentration of IFN-γ ≤225 pg/ml were considered to have pHLH, with a sensitivity of 73% and a specificity of 84%. A four-quadrant model based on the two cutoff values was established to stratify the patients into different clinical situations. The HLH subtypes, cytokine levels, treatment regimens, treatment response, and outcomes were different among the four quadrants, with the 8-week mortality from 2.9 ± 2.9% to 21.4 ± 5.5% and the 5-year overall survival from 93.9 ± 4.2% to 52.6 ± 7.1%. Conclusions: Different subtypes of HLH present distinct cytokine patterns. IFN-γ and the ratio of IL-10 to IFN-γ are helpful tools to differentiate HLH subtypes. A four-quadrant model based on these two parameters is a useful tool for a simple evaluation of the HLH situation.

Clinical features and prognostic impact of TCF3-PBX1 in childhood acute lymphoblastic leukemia: A single-center retrospective study of 837 patients from China.

OBJECTIVES: Conflicting data have been published regarding the prognostic impact of the t(1;19)/TCF3-PBX1 translocation in childhood ALL. The objective of this study was to explore the correlation between the TCF3-PBX1 fusion gene and clinical outcome in Chinese children with newly diagnosed ALL. METHODS: In order to address this issue in our setting, we summarized and analyzed the data of 837 Chinese children with ALL diagnosed between 2010 and 2017. All the patients were treated with the National Protocol of Childhood Leukemia in China (NPCLC)-ALL-2008 protocol. Clinical characteristics and prognosis of pediatric ALL patients with or without TCF3-PBX1 rearrangement were analyzed and compared retrospectively. RESULTS: The TCF3-PBX1 fusion gene was identified in 48 (5.7%) of 837 children with ALL. Our results showed that TCF3-PBX1 positive patients had higher pretreatment white blood cell counts, higher PB blasts percentages and worse risk classification at diagnosis. No statistically significant differences in CR rates, response to prednisone and relapse rates were found between TCF3-PBX1-positive and -negative patients. The 5-year predicted EFS, RFS, and OS of the TCF3-PBX1 positive group compared with the control group were 86.2%±5.3% vs 85.4%±1.3% (P=0.657), 88.2%±5.1% vs 92.2%±1.0% (P=0.458) and 90.4%±4.6% vs 89.0%±1.1% (P=0.561), respectively. No differences were observed regarding clinical outcome between these two groups. When compared with standard risk, intermediate risk and high risk group patients, the long-term survival of TCF3/PBX1 positive group was approximately similar to that of the intermediate risk group under the same protocol in our single center. CONCLUSION: In contrast to previous studies, childhood ALL patients with TCF3-PBX1 transcripts do not appear to show a better outcome than their negative counterparts. TCF3/PBX1 positive was a definitive intermediate risk factor with our NPCLC-ALL-2008 protocol.

Pediatric blastic plasmacytoid dendritic cell neoplasm: report of four cases and review of literature.

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare and aggressive hematological malignancy with poor outcome. Four children with BPDCN treated at our hospital were enrolled. All the four cases presented with cutaneous lesions. Bone marrow and central nervous system was involved in 50% and 25% of patients, respectively. The whole exome sequencing analysis revealed that KMT2 family genes were the most frequently mutated (4/4, 100%), followed by IKZF2 (2/4, 50%). The point mutation p.D348N was found in three patients and one patient had p.C394Y mutation in the KMT2C gene. Translocation of KMT2A-MLLT3 was found in Case 2. Case 1 had complex karyotype, who was induced by acute myeloid leukemia-like regimens. Although he received allogeneic hematopoietic stem cell transplantation twice as well as CD123 chimeric antigen receptor T cell therapy, the disease still progressed and he died 37 months after diagnosis. The other three patients were treated with Interfant-99 protocol. They tolerated the therapy well without significant toxicities and now in complete remission so far with a median follow up time of 9 months. More studies are needed to address the question whether the complex karyotype and KMT2 family genes are the causes of the relapse and refractory in BPDCN.

[Effect of OCT4 Gene Expression Upregulation on the Expression of iPSC-related Transcription Factors in Human Bone Marrow Derived Mesenchymal Stem Cells].

OBJECTIVE: To explore the effect of OCT4 over-expression on the expression of induced pluripotent stem cell (iPSC)-related transcription factors (cMYC，KLF4，LIN28，NANOG and SOX2) in human bone marrow derived mesenchymal stem cells (hBMMSCs), so as to provide fundamental basis for exploring the pathogenesis of hematological diseases (leukemia, aplastic anemia, etc.) from the perspective of hemopoietic microenvironment in the future. METHODS: Recombinant plasmid pcDNA3.1-OCT4 was constructed and transferred into the optimal generation P3-4 hBMMSCs by liposome transfection. The cells with stable and high expression of OCT4（hBMMSCs-OCT4）were screened by G418 resistance screening (limited dilution) and subcloning, the expression of OCT4 mRNA and OCT4 protein was verified by RT-PCR and FCM, respectively. The expression of iPSC-related transcription factors (cMYC, KLF4, LIN28, NANOG and SOX2) were also determined by FCM and RT-PCR, so as to evaluate the effect of ectopic high expression of OCT4 on the expression of iPSC related transcription factors in hBMMSCs. RESULTS: Recombinant plasmid pcDNA3.1-OCT4 was successfully constructed and cells with stable and high expression of OCT4 were successfully screened from hBMMSCs by limited dilution and subcloning. The result of flow cytometry showed that the mean expression level of OCT4 protein increased from (3.03±1.49)% to (95.46±1.40)% compared with the untransfected parental MSCs, which was also confirmed by RT-PCR analysis. At the same time, the expression levels of OCT4 protein and mRNA were compared between transient transfection (day 4) and stable expression cells (day 96), respectively, it was showed that the OCT4 protein level increased from (36.36±0.28)% at day 4 to (96.25±1.38)% at day 96, and the OCT4 mRNA level increased from 2.75-folds to 6.23-folds, respectively. Compared with the untransfected parental MSCs, the average expression levels of stemness transcription factors increased from (1.12±0.47)% (cMYC), (0.84±0.30)% (KLF4), (2.14±0.79)% (LIN28), (0.63±0.37)% (NANOG) and (14.34±2.44)% (SOX2) to (80.65±4.75)%, (73.03±4.70)%, (68.08±3.05)%, (39.39±1.85)%and (91.45±4.56)% in hBMMSCs-OCT4, respectively, which were consistent with results of RT-PCR analysis. Moreover, the expression levels of NANOG and SOX2 positively correlated with the mean expression of OCT4 (OCT4 vs NANOG: r=0.7802，OCT4 vs SOX2: r=0.4981；NANOG vs SOX2: r=0.7426). CONCLUSION: Cells with stable and high expression of OCT4 have been successfully established from hBMMSCs. Ectopic high expression of transcription factor OCT4 in hBMMSCs can up-regulate the expression of other iPSC-related transcription factors such as cMYC, KLF4, LIN28, NANOG and SOX2.

[A genetic analysis of children with Epstein-Barr virus-positive hemophagocytic lymphohistiocytosis and its association with T-helper type 1/T-helper type 2 cytokines].

OBJECTIVE: To study the effect of genetic variation on the prognosis of children with Epstein-Barr virus (EBV)-positive hemophagocytic lymphohistiocytosis (HLH) and its association with cytokines. METHODS: A total of 81 EBV-positive HLH children who received the sequencing of related genes were enrolled. According to the results of gene detection, they were divided into a non-mutation group and a mutation group. According to the pattern of gene mutation, the mutation group was further divided into three subgroups: single heterozygous mutation (SHM), double heterozygous mutation (DHM), and homozygous or compound heterozygous mutation (H-CHM). The serum levels of cytokines were measured and their association with HLH gene mutations was analyzed. RESULTS: UNC13D gene mutation had the highest frequency (13/46, 28%). The STXBP2 c.575G>A(p.R192H) and UNC13D c.604C>A(p.L202M) mutations (likely pathogenic) were reported for the first time. The mutation group had a significantly higher level of tumor necrosis factor alpha (TNF-α) than the non-mutation group, while it had a significantly lower level of interferon gamma (IFN-γ) than the non-mutation group (P<0.05). The IL-4 level of the DHM subgroup was higher than that of the non-mutation group, while the IL-4 level of the H-CHM subgroup was lower than that of the DHM group (P<0.0083). The H-CHM subgroup had a significantly lower 1-year overall survival rate than the non-mutation group, the SHM subgroup, and the DHM subgroup (39%±15% vs 85%±6%/86%±7%/91%±9%, P=0.001). CONCLUSIONS: There is a significant reduction in IFN-γ level in the mutation group. Children with homozygous or compound heterozygous mutation tend to have poorer prognosis, while other mutations do not have a significant impact on prognosis.

High CD38 expression in childhood T-cell acute lymphoblastic leukemia is not associated with prognosis.

BACKGROUND: Prognostic factors are not well exploited in childhood T-cell acute lymphoblastic leukemia (T-ALL). OBJECTIVE: The aim of this study was to analyze the prognostic role of CD38 as well as minimal residual disease (MRD) and other biological factors in T-ALL. METHODS: Immunophenotyping of bone marrow (BM) at diagnosis and MRD levels were determined using a standard panel of antibodies by 4-colour flow cytometry. A total of 96 children with T-ALL were enrolled. RESULTS: The results showed that 97.9% of T-ALL patients were positive for CD38 with a median level of 85.3%. CD38-high group had a worse early treatment response than the CD38-low group. However, CD38 levels were not associated with prognosis, albeit CD38-high group had a worse 5-year event free survival rate (55.1% vs. 66.6%, P> 0.05) and a higher 5-year cumulative incidence of relapse (35.6% vs. 19.8%, P> 0.05). Very high MRD levels (> 10%) were related to the worse survival. Neither flow cytometry based minimal residual disease (MRD) levels nor CD38 expression levels showed significant relation to the hazard of relapse (P> 0.05). CONCLUSIONS: We conclude that T-ALL has a high level of CD38 expression which is not associated with prognosis. Very high MRD level (> 10%) is related to the worse survival, however, FCM based MRD detection does not convey a significant prognostic value.

Dilemmas in diagnosis and management of hemophagocytic lymphohistiocytosis in children.

BACKGROUND: Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening entity which is characterized by severe hyperinflammation. Now the HLH-2004 protocol has been widely accepted and clinically used; however, many questions still remain in clinical practice. In this review, we discuss the dilemmas in the diagnosis and treatment of HLH in children. DATA SOURCES: Original research for articles and literature reviews published in PubMed was carried out using the key term "hemophagocytic lymphohistiocytosis". RESULTS: As the gene sequencing technology progresses, the range of causal mutations and primary HLH has been redefined. The monoallelic variants may contribute to the pathogenesis of the disease. Many conditions without defective cytotoxicity of T or NK cells may lead to HLH, such as primary immunodeficiency (PID) and dysregulated immune activation or proliferation (DIAP). HLH shares overlapping clinical and laboratory characteristics with severe sepsis, but usually the single values are more pronounced in HLH than sepsis. H score is another approach to help the diagnosis of secondary HLH. Specific Th1/Th2 cytokine patterns are very helpful tools to differentiate HLH (reactivation of HLH) from sepsis. Moreover, it also has been used successfully to stratify the therapy intensity. The treatment of HLH should consider underlying diseases, triggers and severity. HLH-94 is recommended for patients who need etoposide-based therapy. CONCLUSIONS: Dramatic progress has been made during the past decades in understanding the pathophysiology of HLH. However, diagnosis and treatment of HLH remain with many dilemmas because of the heterogeneous nature of the disease. Better understanding new gene defects and more effective diagnostic approaches and salvage regimens are goals for the future.

Low expression of TET2 gene in pediatric acute lymphoblastic leukemia is associated with poor clinical outcome.

INTRODUCTION: TET2, a member of the Ten-Eleven translocation gene family, catalyzes the conversion of 5-methylcytosine to 5-hydroxymethylcytosine in DNA. Low expression of TET2 has been reported as a prognostic factor for several types of malignancies in adult patients. However, there have been few data on the effect of TET2 mRNA level on the prognosis of children with ALL so far. METHODS: In this study, TET2 expression of samples cryopreserved in the liquid nitrogen from January 1, 2007 through December 31, 2011 was retrospectively analyzed in 136 newly diagnosed ALL patients by real-time polymerase chain reaction (PCR) assay. The patients' samples were divided into two groups by the median value of patients group and divided into TET2 low and TET2 high groups. RESULTS: A total of 136 childhood ALL patients demonstrated lower TET2 expression than control group (P = .038). TET2 mRNA expression levels were correlated with the disease status. In addition, patients with low TET2 expression had lower platelet counts and lower CR rates. Survival analysis showed that low TET2 expression in children with ALL was associated with lower 5-year overall survival (OS) (63% vs 88%, P = .011) and event-free survival (EFS) (60% vs 85%, P = .003). Multivariate analysis revealed that low TET2 expression was an independent poor prognostic factor of OS and EFS. CONCLUSION: Low expression of TET2 in children with ALL is associated with poor prognosis and can be used as a molecular prognostic marker for risk group stratification.