Genomic signatures and prognosis of advanced stage Chinese pediatric T cell lymphoblastic lymphoma by whole exome sequencing.

Objective: To investigate the genomic signatures and prognosis of advanced-stage T cell lymphoblastic lymphoma (T-LBL) and to examine the relationship between T-LBL and T cell acute lymphoblastic leukemia (T-ALL). Methods: 35 Chinese T-LBL children with stage III or IV disease were recruited for this study. They were treated with combination chemotherapy and whole exome sequencing. The relationship of the clinical features, prognosis and specific gene mutations was researched. Gene chips of T-LBL and T-ALL were downloaded from a database, and differential gene expression was analyzed. Results: Germline causal gene mutations (CARS or MAP2K2) were detected in 2 patients; 3.06 ± 2.21 somatic causal gene mutations were identified in the 35 patients, and somatic mutations were observed in the NOTCH1, FBXW7, PHF6 and JAK3 genes. NOTCH1 mutations were significantly associated with FBXW7 mutations, and the age at diagnosis of patients with NOTCH1-FBXW7 mutations was less than that of patients without such mutations (P < 0.05). 32 patients achieved complete remission (CR), and 14 and 18 patients were classified into the intermediate risk (IR) group and high risk (HR) group. During a median follow-up of 44 months, 3 patients relapsed. Three-year prospective event free survival (pEFS) was 82.286%, and no significant differences of pEFS were found for different sexes, ages, or statuses of NOTCH1-FBXW7 mutations, (P > 0.05); however, the mean survival time of the IR group was longer than that of the HR group (P < 0.05). Differential expression of genes in the T-LBL and/or T-ALL datasets was analyzed using the R package limma, and 1/3 of the differentially expressed genes were found in both the T-ALL and T-LBL datasets. High expression of PI3K-Akt signal pathway genes and the USP34 gene was found in the T-LBL dataset. Conclusion: Although T-ALL and T-LBL both originate from precursor T-cells and are considered different manifestations of the same disease and the outcome of T-LBL is favorable when using T-ALL-based chemotherapy, there are differences in the gene distribution between T-LBL and T-ALL. It seems that the PI3K-Akt signaling pathway and the USP34 gene play important roles in T-LBL, but medicines targeting the USP34 gene or the PI3K-Akt pathway may be invalid.

A study protocol for investigating the sonographic characteristics of neonates with critical illness: an observational cohort study.

BACKGROUND: Haemodynamic instability and hypoxaemia are common and serious threats to the survival of neonates. A growing body of literature indicates that critical care ultrasound has become the optimal evaluation tool for sick neonates. However, few studies have described sonographic characteristics of haemodynamics systematically in the neonates with critical illness. This protocol describes a prospective observational cohort study aimed at (1) characterising the sonographic characteristics of the neonates with critical diseases; and (2) assessing the mortality, significant morbidity, utility of vasoactive medications, fluid resuscitation, duration of ventilation, etc. METHODS AND ANALYSIS: This is a single-centre, prospective and observational study conducted in Chengdu Women's and Children's Central Hospital from 1 December 2022 to 31 December 2027. Neonates admitted to the neonatal intensive care unit will be recruited. After inclusion, the neonates will undergo the neonatal critical care ultrasound. The data collected via case report forms include clinical variables and sonographic measures. The primary outcome is to identify the sonographic characteristics of sick neonates with different diseases, and the secondary outcome is to describe the mortality, significant morbidity, utility of vasoactive medications, fluid resuscitation and duration of ventilation. DISCUSSION: Our study provided an organised neonatal critical care ultrasound workflow, which can be applied in practice. Accordingly, this study will first set up large data on the sonographic description of the neonates with critical illness, which can help to understand the pathophysiology of the critical illness, potentially titrating the treatment. TRIAL REGISTRATION NUMBER: Chinese Clinical Trial Registry (ChiCTR2200065581; https://www.chictr.org.cn/com/25/showproj.aspx?proj=184095).

[The Application of RNA-Sequencing in Pediatric B-Cell Acute Lymphoblastic Leukemia].

OBJECTIVE: To explore the molecular genetic characteristics of children with B-cell acute lymphoblastic leukemia (B-ALL) and the application value of RNA-sequencing (RNA-seq). METHODS: The clinical and laboratory examination data of newly diagnosed B-ALL children who were given treatment in the Department of Hematology, Children's Hospital of Chongqing Medical University from May 2015 to April 2020 were collected and analyzed. All children were confirmed by bone marrow morphology, histochemical staining and flow cytometry, and the karyotype analysis, FISH, RT-PCR and RNA-seq detection were conducted. RESULTS: There were 71 males and 58 females with a median age of 50(8-190) months in 129 newly diagnosed children with B-ALL. The fusion gene was positive in 99 children (76.7%). A total of 86 leukemia related or possibly related gene mutations were detected, with a positive rate of 66.7%. There was no significant difference in the detection rates of ETV6-RUNX1, BCR-ABL1, TCF3-PBX1 and KMT2A rearrangements among FISH, RT-PCR and RNA-seq. Rare fusion genes were detected by RNA-seq, including 1 case of KMT2A-USP2, 4 cases of Ph-like related fusion genes, 5 cases of MEF2D rearrangement, 5 cases of PAX5 rearrangement, 3 cases of ZNF384 rearrangement, as well as several fusion genes whose significance were not clear or had not been reported in children with leukemia. Besides, children with ETV6-RUNX1 fusion gene had good response to induction of remission, while children with BCR-ABL1 and ZNF384 rearrangement had poor response, the remission rate of minimal residual disease was statistically significant compared with other types (P<0.05). CONCLUSION: RNA-seq can not only detect known fusion genes, but also discover new or rare fusion genes and gene mutations. The application of RNA-seq has important guiding significance for risk classification and precise targeted therapy of pediatric B-ALL.

The long-term outcome and risk factors for precursor B cell acute lymphoblastic leukemia without specific fusion genes in Chinese children: experiences from multiple centers.

Specific fusion genes play important roles as risk factors for strategic treatment in pediatric B-cell acute lymphoblastic leukemia (B-ALL), and the risk factors in patients without common fusion genes have not been well demonstrated. We collected and analyzed clinical and laboratory findings, treatment responses and outcomes in B-ALL patients without specific fusion genes. Whole-exome sequencing (WES) and/or RNA sequencing (RNAseq) data from bone marrow relapsed patients were also analyzed. 283 patients were enrolled in the study. Traditional elements and treatment responses at different time points (TPs) were evaluated to classify risk groups and adjust the treatment strategy. Treatment-related mortality was found in 11 (3.89%) patients, 49 (17.31%) patients relapsed, and the ten-year prospective event-free survival (pEFS) was 78.2±2.5%. Univariate analysis revealed that significant differences were not found in the pEFS of traditional risk factors, including sex, age, WBC count or chromosome status; good responses of BM smears at TP1 and minimal residual disease (MRD) levels at TP2 and TP3 were strongly associated with prolonged pEFS. Compared with the IR or the HR group, patients in the SR group presented with longer pEFS and a lower relapse rate. Multivariable analysis of outcomes and hazard ratios revealed that a positive MRD level was a key risk factor. WES or RNAseq was performed for BM relapse patients, and adverse and unreported genetic abnormalities were discovered. Favorable outcomes were acquired in the cohort. The study results showed that traditional risk factors and poor prednisone response were overcome by modified chemotherapy, and a positive MRD level was a key risk factor in these patients. NGS is needed to discover more risk-related molecular abnormalities.

[Second-generation Sequencing Analysis of Ph+ and Ph-like Childhood T-cell Acute Lymphoblastic Leukemia].

OBJECTIVE: To screen the core genes of Philadelphia chromosome positive/Ph like T-cell acute lymphoblastic leukemia (Ph+/Ph like T-ALL) using bioinformatics methods, and analyze the core sub-networks for exploration of the development process of Ph+/Ph like T-ALL, so as to find the molecular targets that may be used in clinical diagnosis and treatment. METHODS: The WES/RNA-seq examination results of Ph+/Ph-like T-ALL children had be collected in our hospital, the genetic data that met the requirements had be downloaded from GEO database, then GRO2R online differentially expressed gene screening program was used to screen differentially expressed genes, finally, GO function enrichment analysis and KEGG pathway enrichment analysis were performed to compare differentially expressed genes. At the same time, STRING database and Cytoscape software were used to build protein interaction network and screen hub genes and core sub-networks. RESULTS: For Ph+/Ph like T-ALL, a total of 84 differentially expressed genes had been found, for Ph+ ALL a total of 249 differentially expressed genes, and for T-ALL a total of 175 differentially expressed genes. Based on the results of GO function enrichment, KEGG pathway enrichment analysis and protein interaction network, RPA1, POLD1, POLE and SOCS1 were selected as hub genes. DNA damage repair and JAK/STAT signal transduction pathway were the main functional sub-networks. CONCLUSION: There are obviously abnormal DNA damage repair pathways in children with Ph+/Ph like T-ALL. RPA1, POLD1 and POLE may be important relevant biomarkers for the occurrence and development of Ph+/Ph like T-ALL, which can provide a basis for further research.

Acute myeloid leukemia arising to genetic susceptibility genes related T cell acute lymphoblastic leukemia: case report.

Fanconi anemia (FA) is the most common inherited bone marrow failure disorder, with a predisposition to neoplasia. While Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are the most common hematologic malignancies seen in patients with FA, cases of acute lymphoblastic leukemia (ALL) have also been described in the literature but it is uncommon. In our case report, a 12 years 5 months old boy, who was detected with heterozygote mutation of FANCC gene and nonsynonymous single nucleotide variability (SNV) mutation of AKAP9 gene, presented with precursor T cell ALL (T-ALL) at onset, myelodysplasia or myeloid biomarkers were not found at initial diagnosis. He received chemotherapy and achieved complete remission (CR) after a course of remission induction, but severe cytopenia was presented, sepsis and Invasive fungal infection also arose. With following-up and continue chemotherapy, secondary AML arose 17 months later, the patient died of sepsis related to chemotherapy at AML status. FA patients usually presented with homozygous or bilateral heterozygosity mutation in literature reports, whereas heterozygosity gene mutation of FANCC and AKAP9 has not reported yet. AKAP9 protein which was encoded by AKAP9 gene is widely distributed in many kinds of cells, thus ensuring the specificity and accuracy of signal transduction. We speculate that AKAP9 protein may interfere with the normal signal transduction of heterozygous mutation expression of FANCC gene and result in the inactivation of FANCC gene function. Unfortunately, the patient died of sepsis and we don't have enough blood samples to explore the role of AKAP9 gene mutation in patients with heterozygosity FANCC gene mutation.

Mature B cell acute lymphoblastic leukaemia with KMT2A-MLLT3 transcripts in children: three case reports and literature reviews.

BACKGROUND: Mature B cell acute lymphoblastic leukaemia (BAL) is characterised by French-American-British (FAB)-L3 morphology and the presence of surface immunoglobulin (sIgM) light chain restriction. BAL is also considered as the leukaemic phase of Burkitt lymphoma (BL), in which t (8; 14) (q24; q32) or its variants are related to the myelocytomatosis oncogene (MYC) rearrangement (MYCr) is usually present. However, BAL with lysine methyltransferase 2A (KMT2A, previously called Mixed lineage leukaemia, MLL) gene rearrangement (KMT2Ar, previously called MLLr) is rare. RESULTS: Three BAL patients with KMT2Ar were enrolled between January 2017 and November 2019, accounting for 1.37% of the B-ALL population in our hospital. We also reviewed 24 previously reported cases of BAL and KMT2Ar and analysed the features, treatment, and prognosis. Total 13 males and 14 females were enrolled in our research, and the average age at diagnosis was 19.5 ± 4.95 months old. In these 27 patients, renal, central nervous system (CNS) and skin involvement were existent in 6, 4 and 3 patients, respectively; 26 patients (26/27) showed non-ALL-L3 morphology, while one patient is ALL-L3; overexpression of CD19 was detected in most cases, negative or suspicious expression of CD20 was found in 64% of patients. KMT2Ar was reported, but MYCr was not observed. 25 patients (25/27) achieved complete remission after chemotherapy or Stem cell transplantation. The patients were sensitive to chemotherapy, prospective event-free survival (pEFS) of BAL patients with KMT2Ar who received allogeneic haematopoietic stem cell transplantation (allo-HSCT) was higher than that in patients who received chemotherapy alone (83.33% vs 41.91%). CONCLUSION: BAL patients with KMT2Ar had unique manifestations, including younger age at diagnosis and overexpression of CD19; expression of CD20 was rare, and MYCr was undetectable. The pEFS was higher in patients undergoing allo-HSCT than in patients undergoing chemotherapy alone.