Preclinical evaluation of cyclophosphamide and fludarabine combined with CD19 CAR-T in the treatment of B-cell hematologic malignancies in vivo.

Background: Chimeric antigen receptor T (CAR-T) cell therapy has achieved marked therapeutic success in ameliorating hematological malignancies. However, there is an extant void in the clinical guidelines concerning the most effective chemotherapy regimen prior to chimeric antigen receptor T (CAR-T) cell therapy, as well as the optimal timing for CAR-T cell infusion post-chemotherapy. Materials and Methods: We employed cell-derived tumor xenograft (CDX) murine models to delineate the optimal pre-conditioning chemotherapy regimen and timing for CAR-T cell treatment. Furthermore, transcriptome sequencing was implemented to identify the therapeutic targets and elucidate the underlying mechanisms governing the treatment regimen. Results: Our preclinical in vivo evaluation determined that a combination of cyclophosphamide and fludarabine, followed by the infusion of CD19 CAR-T cells five days subsequent to the chemotherapy, exerts the most efficacious therapeutic effect in B-cell hematological malignancies. Concurrently, RNA-seq data indicated that the therapeutic efficacy predominantly perturbs tumor cell metabolism, primarily through the inhibition of key mitochondrial targets, such as C-Jun Kinase enzyme (C-JUN). Conclusion: In summary, the present study offers critical clinical guidance and serves as an authoritative reference for the deployment of CD19 CAR-T cell therapy in the treatment of B-cell hematological malignancies.

miR-133a and miR-135a Regulate All-Trans Retinoic Acid-Mediated Differentiation in Pediatric Acute Myeloid Leukemia by Inhibiting CDX2 Translation and Serve as Prognostic Biomarkers.

Background: Acute myeloid leukemia (AML) is a type of blood cancer characterized by excessive growth of immature myeloid cells. Unfortunately, the prognosis of pediatric AML remains unfavorable. It is imperative to further our understanding of the mechanisms underlying leukemogenesis and explore innovative therapeutic approaches to enhance overall disease outcomes for patients with this condition. Methods: Quantitative reverse-transcription PCR was used to quantify the expression levels of microRNA (miR)-133a and miR-135a in 68 samples from 59 pediatric patients with AML. Dual-luciferase reporter transfection assay, Cell Counting Kit-8 assay, and western blot analysis were used to investigate the functions of miR-133a and miR-135a. Results: Our study found that all-trans-retinoic acid (ATRA) promoted the expression of miR-133a and miR-135a in AML cells, inhibited caudal type homeobox 2 (CDX2) expression, and subsequently inhibited the proliferation of AML cells. Additionally, miR-133a and miR-135a were highly expressed in patients with complete remission and those with better survival. Conclusions: miR-133a and miR-135a may play an antioncogenic role in pediatric AML through the ATRA-miRNA133a/135a-CDX2 pathway. They hold promise as potentially favorable prognostic indicators and novel therapeutic targets for pediatric AML.

Identification of the Shared Gene Signatures of HCK, NOG, RNF125 and Biological Mechanism in Pediatric Acute Lymphoblastic Leukaemia and Pediatric Sepsis.

The shared mechanisms between pediatric acute lymphoblastic leukaemia (ALL) and pediatric sepsis are currently unclear. This study was aimed to explore the shared key genes of pediatric ALL and pediatric sepsis. The datasets involved were downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) between disease and control samples in GSE13904 and GSE79533 were intersected. The least absolute shrinkage and selection operator (LASSO) and the boruta analyses were performed in GSE13904 and GSE79533 separately based on shared DEGs, and shared key genes were obtained by taking the intersection of sepsis-related key genes and ALL-related key genes. Three shared key genes (HCK, NOG, RNF125) were obtained, that have a good diagnostic value for both sepsis and ALL. The correlation between shared key genes and differentially expressed immune cells was higher in GSE13904 and conversely, the correlation of which was lower in GSE79533. Suggesting that the sharing key genes had a different impact on the immune environment in pediatric ALL and pediatric sepsis. We make the case that this study provides a new perspective to study the relationship between pediatric ALL and pediatric sepsis.

Lycorine eliminates B-cell acute lymphoblastic leukemia cells by targeting PSAT1 through the serine/glycine metabolic pathway.

B-cell acute lymphoblastic leukemia (B-ALL) has been confirmed as the most common malignant hematologic neoplasm among children. A novel antitumor mechanism of lycorine was elucidated in this study. As revealed by the result of this study, lycorine significantly inhibited the growth and proliferation of REH and NALM-6 and induced their apoptosis. The result of the RNA-seq analysis suggested that lycorine targeted PSAT1 of serine/glycine metabolism in B-ALL cells. As indicated by the result of the GSEA analysis, the genes enriched in the amino acid metabolic pathways were down-regulated by lycorine. As revealed by the results of ectopic expression, shRNA knockdown assays, and further liquid-phase tandem mass spectrometry (LC-MS) analysis, lycorine reduced serine/glycine metabolites by down-regulating PSAT1, further disrupting carbon metabolism and eliminating B-ALL cells. Furthermore, lycorine showed a synergistic effect with cytarabine in ALL treatments. Lastly, lycorine significantly down-regulated leukemia progression in the cell line-derived xenograft (CDX) model. In brief, this study has suggested for the first time that lycorine is a promising anti-ALL drug, and a novel amino acid metabolism-associated property of lycorine was identified.

Clinical efficacy analysis of paxlovid in children with hematological diseases infected with the omicron SARS-CoV-2 new variant.

Objective: To summarize the clinical characteristics of children with hematological malignancies co-infected with novel coronavirus and explore the safety and effectiveness of Paxlovid treatment. Methods: From December 10, 2022, to January 20, 2023, the clinical data of children with hematological diseases diagnosed with novel coronavirus infection in the outpatient and emergency department of the Seventh Affiliated Hospital of Sun Yat-sen University were retrospectively analyzed. Results: According to whether to give paxlovid or not, it is divided into group A (paxlovid group) and group B (non-paxlovid group). The length of fever was 1-6 days in group A and 0-3 days in group B. The viral clearance time was shorter in group A than in group B. The inflammatory indexes CRP and PCT were significantly higher in group A than in group B (P < 0.05). Twenty patients were followed up for 1 month after leaving the hospital, and there were 5 cases of reappearance of fever, 1 case of increased sleep, 1 case of physical fatigue and 1 case of loss of appetite within 2 weeks. Conclusions: Paxlovid has no apparent adverse reactions in children 12 years old and younger with underlying hematological diseases infected with the new coronavirus. Focusing on the interaction between paxlovid and other drugs is necessary during the treatment.

Comprehensive analysis of PTPN family expression and prognosis in acute myeloid leukemia.

Background: Tyrosyl phosphorylation is carried out by a group of enzymes known as non-receptor protein tyrosine phosphatases (PTPNs). In the current investigation, it is hoped to shed light on the relationships between the expression patterns of PTPN family members and the prognosis of acute myeloid leukemia (AML). Methods: PTPN expression was examined using GEPIA and GEO databases. To investigate the connection between PTPN expression and survival in AML patients, we downloaded data from the Broad TCGA Firehose and Clinical Proteomic Tumor Analysis (CPTAC) of the Cancer Genome Atlas (TCGA). We used quantitative real-time PCR (qRT-PCR) to confirm that essential genes were performed in clinical samples and cell lines. We then used western blot to verify that the genes expressed in the above databases were positive in normal tissues, AML patient samples, and AML cell lines. Next, we investigated associations between genome-wide expression profiles and PTPN6 expression using the GEO datasets. We investigated the interactive exploration of multidimensional cancer genomics using the cBioPortal datasets. Using the DAVID database, a study of gene ontology enrichment was performed. The protein-protein interaction (PPI) network was created using the STRING portal, and the gene-gene interaction network was performed using GeneMANIA. Results: Data from GEO and GEPIA revealed that most PTPN family members were linked to AML. Patients with leukemia have elevated levels of several PTPN members. All of the AML patients' poor overall survival (OS, p < .05) was significantly linked with higher expression of PTPN1, PTPN6, and PTPN7. Additionally, clinical samples showed that the expression of PTPN 6, PTPN 7, PTPN 13, and PTPN 14 was higher than normal in AML patients (p = .0116, p = .0034, p = .0092, and p = .0057, respectively) and AML cell lines (p = .0004, p = .0035, p = .0357, and p = .0177, respectively). Western blotting results showed that the expression of PTPN6 in AML samples and AML cell lines was significantly higher than that in normal control samples. Conclusion: Differentially expressed PTPN family members were found in AML. The prognosis of patients and PTPN gene expression were shown to be correlated. PTPN6 is one of these members and may be used as an AML diagnostic and prognostic marker.

Clinical Outcome of Acquired Post-Immunosuppressive-Therapy Aplastic Anemia in Pediatric Patients: A 13-Year Experience in Two Southern China Tertiary Care Centers.

OBJECTIVE: The aim of the present study is to evaluate the efficacy, complications, and contributing factors of immunosuppressive therapy (IST) response in children with acquired aplastic anemia (AA) and to explore optimal therapeutic methods for different clinical AA types. METHODS: A total of 130 children diagnosed with acquired AA underwent IST in the Department of Pediatrics at Sun Yat-sen Memorial Hospital and the Department of Pediatrics at Seventh Affiliated Hospital, Sun Yat-sen University, between January 1, 2006, and July 15, 2020. The overall survival (OS), response rates, complications, and response predictors were analyzed. The response rates were compared according to clinical AA type. RESULTS: All 130 children with AA were followed up with for a median of 50.6 months. Among the patients, 25 had non-severe AA (NSAA), 64 had severe AA (SAA), and 41 had very severe AA (VSAA). All patients initially received IST. In 13 patients, the IST failed; these patients received an allo-hematopoietic stem cell transplant as a salvage regimen. The OS rate was 90.3% ± 2.8%, and the response rates at 3, 6, 9, and 12 months were 34.19%, 39.32%, 49.57%, and 66.67%, respectively. The prolonged follow-up period might have led to higher response rates, especially in patients with SAA and VSAA. A multivariate logistic regression analysis of prognostic factors was conducted; the results showed that high red blood cell (RBC) and platelet (PLT) counts were associated with a high overall response rate and that the RBC count at diagnosis is a major contributing factor. CONCLUSION: With the use of rabbit anti-thymocyte globulin, proper cyclosporine management, and a prolonged IST follow-up period, a higher number of patients with acquired AA than normal achieved response. Proportionally, the number of patients who achieved remission within 12 months was higher in the SAA group (38.18%→63.64%) and VSAA group (28.95%→65.79%) than in the NSAA group (58.33%→75%). Higher RBC and PLT counts at diagnosis can predict a favorable outcome.

Performance of PRISM III, PELOD-2, and P-MODS Scores in Two Pediatric Intensive Care Units in China.

Objective: The performances of the pediatric risk of mortality score III (PRISM III), pediatric logistic organ dysfunction score-2 (PELOD-2), and pediatric multiple organ dysfunction score (P-MODS) in Chinese patients are unclear. This study aimed to assess the performances of these scores in predicting mortality in critically ill pediatric patients. Methods: This retrospective observational study was conducted at two tertiary-care PICUs of teaching hospitals in China. A total of 1,253 critically ill pediatric patients admitted to the two Pediatric Intensive Care Units (PICUs) of the First Affiliated Hospital, Sun Yat-Sen University from August 2014 to December 2019 and Shen-Zhen Children's Hospital from January 2019 to December 2019 were analyzed. The indexes of discrimination and calibration were applied to evaluate score performance for the three models (PRISM III, PELOD-2, and P-MODS scores). The receiver operating characteristic (ROC) curve was plotted, and the efficiency of PRISM III, PELOD-2, and P-MODS in predicting death were evaluated by the area under ROC curve (AUC). Hosmer-Lemeshow goodness-of-fit test was used to evaluate the degree of fitting between the mortality predictions of each scoring system and the actual mortality. Results: A total of 1,253 pediatric patients were eventually enrolled in this study (median age, 38 months; overall mortality rate, 8.9%; median length of PICU stay, 8 days). Compared to the survival group, the non-survival group showed significantly higher PRISM III, PELOD-2, and P-MODS scores [PRISM III: 18 (12, 23) vs. 11 (0, 16); PELOD-2, 8 (4, 10) vs. 4 (0, 6); and P-MODS: 5 (4, 9) vs. 3 (0, 4), all P < 0.001]. ROC curve analysis showed that the AUCs of PRISM III, PELOD-2, and P-MODS for predicting the death of critically ill children were 0.858, 0.721, and 0.596, respectively. Furthermore, in the Hosmer-Lemeshow goodness-of-fit test, PRISM III and PELOD-2 showed the better calibration between predicted mortality and observed mortality (PRISM III: χ2 = 5.667, P = 0.368; PELOD-2: χ2 = 9.582, P = 0.276; P-MODS: χ2 = 12.449, P = 0.015). Conclusions: PRISM III and PELOD-2 can discriminate well between survivors and non-survivors. PRISM III and PELOD-2 showed the better calibration between predicted and observed mortality, while P-MODS showed poor calibration.

Astragalus polysaccharide has a protective effect on hematopoiesis in an irradiated mouse model and decreases apoptosis in megakaryocytes.

Huangqi, the dried root of Radix Astragali, is an essential herb in Traditional Chinese Medicine and has been used to promote hematopoiesis for centuries. Astragalus polysaccharide (ASPS), the bioactive compound of Huangqi, serves a crucial role in hematopoiesis. The aim of the present study was to investigate the hematopoietic effects, in particular the thrombopoietic effects, and the molecular mechanisms of ASPS using an irradiation­induced myelosuppressive mouse model. Colony­forming unit assays, flow cytometric analysis of apoptosis, ELISAs, Giemsa staining and western blotting were performed to determine the hematopoietic and anti­apoptotic effects of ASPS. The results demonstrated that ASPS enhanced the recovery of red blood cells at day 21 following treatment, as well as platelets and white blood cells at day 14. In addition, ASPS promoted colony formation in all lineages (megakaryocytes, granulocyte monocytes, erythroid cells and fibroblasts). The morphological study of the bone marrow demonstrated that tri­lineage hematopoiesis was preserved in the ASPS­ and thrombopoietin (TPO)­treated groups compared with the control group. The overall cellularity (mean total cell count/area) of the ASPS­treated group was similar to that of the TPO­treated group. Additionally, in vitro experiments indicated that treatment with 100 µg/ml ASPS exhibited the maximum effect on colony formation. ASPS attenuated cell apoptosis in megakaryocytic cells via inhibiting the mitochondrial caspase­3 signaling pathway. In conclusion, ASPS promoted hematopoiesis in irradiated myelosuppressive mice possibly via enhancing hematopoietic stem/progenitor cell proliferation and inhibiting megakaryocytes apoptosis.

Clinicopathological features, risk and survival in lung cancer survivors with therapy-related acute myeloid leukaemia.

BACKGROUND: A secondary malignancy is the most serious complication in lung cancer (LC) survivors. This study aimed to evaluate the clinicopathological features, predictable risk factors and survival of patients with LC who developed therapy-related acute myeloid leukaemia (t-AML). METHODS: Patients from the Surveillance, Epidemiology, and End Results (SEER) database diagnosed with t-AML after LC between 1975 and 2015 were included. Standardized incidence ratios (SIRs) were used to perform multiple primary analyses. The risk of t-AML development among LC patients was assessed using a logistic regression model. Kaplan-Meier analysis was used to construct overall survival (OS) curves. Cox regression was used to assess the influence of various prognostic factors. RESULTS: A total of 104 patients with t-AML after LC-targeting chemotherapy were included. The median latency period to the development of t-AML was 35.5 months. The calculated SIR of t-AML was 4.00. Chemoradiotherapy, small cell lung cancer (SCLC), or localized/regional-stage LC was a risk factor for the development of t-AML. The median OS was only 1 month, and those younger than 65 years were predicted to have a better OS time. CONCLUSIONS: t-AML is a rare but serious late complication in LC patients and is associated with a poor prognosis. It is necessary to carry out long-term follow-up and screen for t-AML in LC patients, especially among those undergoing both radiotherapy and chemotherapy, with SCLC or with localized/regional-stage LC.

Posaconazole aggravates vincristine-related hypertension in children with acute lymphoblastic leukemia: a case report.

Vincristine-related secondary hypertension is rare. This study reports two children who were treated with vincristine for acute lymphoblastic leukemia (ALL) and posaconazole for fungal infections who experienced vincristine-related secondary hypertension. Blood pressure normalized in both children after halting the drugs and providing antihypertensive treatment. Thus, posaconazole can interact with vincristine and induce secondary hypertension in children with ALL. As an adverse event, this interaction is a rare occurrence.

[Predictive value of four pediatric scores of critical illness and mortality on evaluating mortality risk in pediatric critical patients].

OBJECTIVE: To assess the performance of pediatric clinical illness score (PCIS), pediatric risk of mortality score III (PRISM III), pediatric logistic organ dysfunction score 2 (PELOD-2), and pediatric multiple organ dysfunction score (P-MODS) in predicting mortality in critically ill pediatric patients. METHODS: The data of critically ill pediatric patients admitted to Pediatric Intensive Care Unit (PICU) of First Affiliated Hospital of Sun Yat-Sen University from August 2012 to May 2017 were retrospectively analyzed. The gender, age, basic diseases, the length of PICU stay were collected. The children were divided into survival group and non-survival group according to the clinical outcome during hospitalization. The variables of PCIS, PRISM III, PELOD-2, and P-MODS were collected and scored. Receiver operating characteristic (ROC) curve was plotted, the efficiency of PCIS, PRISM III, PELOD-2, and P-MODS for predicting death were evaluated by the area under ROC curve (AUC). Hosmer-Lemeshow goodness of fit test was used to evaluate the fitting degree of each scoring system to predict the mortality and the actual mortality. RESULTS: Of 461 critically ill children, 35 children were excluded because of serious data loss, hospital stay not exceeding 24 hours, and death within 8 hours after admission. Finally, a total of 426 pediatric patients were enrolled in this study. 355 pediatric patients were survived, while 71 were not survived during hospitalization, with the mortality of 16.7%. There was no significant difference in gender, age, underlying diseases or length of PICU stay between the two groups. PCIS score in non-survival group was significantly lower than that of survival group [80 (76, 88) vs. 86 (80, 92)], and PRISM III, PELOD-2 and P-MODS scores were significantly increased [PRISM III: 16 (13, 22) vs. 12 (10, 15), PELOD-2: 6 (5, 9) vs. 4 (2, 5), P-MODS: 6 (4, 9) vs. 3 (2, 6), all P < 0.01]. ROC curve analysis showed that the AUCs of PCIS, PRISM III, PELOD-2, and P-MODS for predicting death of critical ill children were 0.649, 0.731, 0.773, and 0.747, respectively. Hosmer-Lemeshow test showed that PCIS predicted the mortality and the actual mortality in the best fitting effect (χ 2 = 7.573, P = 0.476), followed by PELOD-2 and P-MODS (χ12 = 9.551, P1 = 0.145; χ22 = 10.343, P2 = 0.111), while PRISM III had poor fitting effect (χ2 = 43.549, P < 0.001). CONCLUSIONS: PRISM III, PELOD-2 and P-MODS can discriminate between survivors and moribund patients well, and assessing the condition of critically ill pediatric patients with relatively accuracy. PCIS was the best fitting effect in predicting mortality and actual mortality, followed by PELOD-2 and P-MODS, while PRISM III had poor fitting effect.

Serum miR-126-3p level is down-regulated in sepsis patients.

BACKGROUND: Endothelial injury is part of the pathogenesis of sepsis. The microRNA-126 (miR-126) was previously identified as an endothelial biomarker and is known to play a critical role in preserving endothelial cell integrity. However, the role of miRNA-126 in sepsis is unclear. METHOD: Blood samples were collected from sepsis patients at the first Affiliated Hospital of Sun Yat-sen University within 24 h (n = 60) and on day 7 (n = 51) after diagnosis, and once from control subjects (n = 46). MiR-126-3p expression was evaluated by quantitative real-time PCR. The miR-126-3p level was correlated with clinical data and a set of routine and experimental biomarkers. The outcome of sepsis patients was determined by follow-up at 28 days after collection of blood samples on day 7. RESULT: MiR-126-3p level was significantly downregulated in sepsis patients 24 h after diagnosis compared with control subjects. Degree of downregulation of serum miR-126-3p correlated with the severity of sepsis. To determine the diagnostic accuracy of miR-126-3p, the receiver operating characteristic (ROC) was performed and the AUC of miR-126-3p was 0.735. Furthermore, serum miR-126-3p concentration at this time point was correlated with the expression markers of systemic inflammation, bacterial infection, and renal and hepatic dysfunction. However, serum miR-126-3p level on day 7 day did not differ between surviving sepsis patients and those who died. CONCLUSION: These results indicate that miR-126-3p could be a diagnostic biomarker for sepsis.

De novo SCN2A mutation in a Chinese infant with severe early-onset epileptic encephalopathy, bronchopulmonary dysplasia, and adrenal hypofunction.

Early-onset epileptic encephalopathies (EOEEs) are a group of phenotypically and genetically heterogeneous neurodevelopmental disorders. Mutations of SCN2A, the gene encoding the aII subunit of the voltage-gated sodium channel (Nav1.2), have been detected in some EOEE patients. This report describes a 4-month-old female who presented with severe EOEE as well as bronchopulmonary dysplasia and adrenal hypofunction. Whole-exome sequencing revealed a novel missense mutation in SCN2A (c.1261T > G; p.L421V) that was not detected in either her parents or her brother. The mutation was confirmed by Sanger sequencing and characterized as pathogenic by several prediction programs. This finding of a de novo SCN2A mutation in an ethnic Chinese infant with EOEE as well as multi-organ dysfunction expands the phenotypic spectrum of SCN2A mutations.

A novel compound heterozygous GAA mutation in a Chinese family with juvenile onset form of Pompe disease with cardiomyopathy.

Pompe disease is an autosomal recessive disorder resulting from a deficiency of acid α-glucosidase (GAA). It is uncommon in the mainland of China, due to rare mutations in the GAA gene. The aim of this work was to elucidate the causative role of a novel compound heterozygous mutation of juvenile onset Pompe disease. In this study, clinical samples were obtained from two siblings with muscle weakness, recurrent airway infections, cardiomyopathy and respiratory insufficiency in a non-consanguineous Chinese family. The α-glucosidase activity in leukocytes of both children was low. Next-generation sequencing was performed on the 19 coding exons of GAA in both children, with confirmation by Sanger sequencing. Next-generation sequencing showed the same compound heterozygous GAA mutation (c.1216G>A p.Asp406Asn and c.1935C>A p.Asp645Glu) in both children. As this mutation is consistent with the clinical manifestations of juvenile onset Pompe disease and no other mutations were detected after scanning the gene sequence, we suggest that the Pompe disease phenotype is caused by compound heterozygosity for c.1216G>A and c.1935C>A. As c.1216G>A is not currently listed in the Pompe disease Mutation Database, this information about Pompe disease in a Chinese population is of particular interest.

A new compound heterozygous CFTR mutation in a Chinese family with cystic fibrosis.

INTRODUCTION: Cystic fibrosis (CF) is the most common autosomal recessive disease among Caucasians but is rarer in the Chinese population, because mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. OBJECTIVES: To elucidate the causative role of a novel compound heterozygous mutation of CF. MATERIALS AND METHODS: In this study, clinical samples were obtained from two siblings with recurrent airway infections, clubbed fingers, salt-sweat and failure to gain weight in a non-consanguineous Chinese family. Next-generation sequencing was performed on the 27 coding exons of CFTR in both children, with confirmation by Sanger sequencing. RESULTS: Next-generation sequencing showed the same compound heterozygous CFTR mutation (c.865A>T p.Arg289X and c.3651_3652insAAAT p.Tyr1219X) in both children. CONCLUSIONS: As this mutation is consistent with the clinical manifestations of CF and no other mutations were detected after scanning the gene sequence, we suggest that the CF phenotype is caused by compound heterozygosity for c.865A>T and c.3651_3652insAAAT. As c865A>T is not currently listed in the "Cystic Fibrosis Mutation Database", this information about CF in a Chinese population is of interest.