A study of the TP53 Germline Mutation and Clinicopathologic Features in Thai Children with Adrenocortical Carcinoma

Objective: To determine the clinicopathologic features and germline tumor protein p53 (TP53) mutation in children with adrenocortical carcinoma (ACC). Material and Methods: This was a retrospective study. From 2009 to 2018, children with ACC from King Chulalongkorn Memorial Hospital and Phramongkutklao Hospital were enrolled into the study. Clinical presentations and hormonal profiles were recorded. Mutation analyses of the TP53 gene were acquired using the next-generation-sequencing method which was performed for germline samples of all patients and the parents of those who tested positive. Result: Two males and six females with ACC were enrolled into this study. The median age at diagnosis was 25.5 months (range 10 to 67 months). All participants had virilization, either virilization only (n=3) or associated with Cushing (n=5). All participants had had surgery to completely remove all of the tumor and three participants had received adjuvant chemotherapy consisting of etoposide, doxorubicin, and cyclophosphamide. All participants had three different known mutations in the TP53 gene: c.1010G>A (p.R337H), c.916C>T (p.R306*) and c.743G>A (p.R248Q). Two of the three participants with TP53 mutations had pulmonary metastasis. One participant had wild-type TP53 and pulmonary recurrence occurred one year after diagnosis. The median follow-up time was 31 months (range 10 to 168 months. As of this writing, seven participants survived without evidence of recurrence. No second malignancy was found in all participants. One participant who had c.916C>T died of pulmonary metastasis 10 months after diagnosis. Conclusion: We successfully identified three different germline TP53 mutations in patients with ACC. Progression to pulmonary metastasis and recurrence were higher among participants with TP53 mutations. The treatment outcome of childhood ACC was good when surgery and adjuvant chemotherapy were used

Role of Mitochondrial Electron Transport Chain Complexes in Capsaicin Mediated Oxidative Stress Leading to Apoptosis in Pancreatic Cancer Cells

We evaluated the mechanism of capsaicin-mediated ROS generation in pancreatic cancer cells. The generation of ROS was about 4–6 fold more as compared to control and as early as 1 h after capsaicin treatment in BxPC-3 and AsPC-1 cells but not in normal HPDE-6 cells. The generation of ROS was inhibited by catalase and EUK-134. To delineate the mechanism of ROS generation, enzymatic activities of mitochondrial complex-I and complex-III were determined in the pure mitochondria. Our results shows that capsaicin inhibits about 2.5–9% and 5–20% of complex-I activity and 8–75% of complex-III activity in BxPC-3 and AsPC-1 cells respectively, which was attenuable by SOD, catalase and EUK-134. On the other hand, capsaicin treatment failed to inhibit complex-I or complex-III activities in normal HPDE-6 cells. The ATP levels were drastically suppressed by capsaicin treatment in both BxPC-3 and AsPC-1 cells and attenuated by catalase or EUK-134. Oxidation of mitochondria-specific cardiolipin was substantially higher in capsaicin treated cells. BxPC-3 derived ρ0 cells, which lack mitochondrial DNA, were completely resistant to capsaicin mediated ROS generation and apoptosis. Our results reveal that the release of cytochrome c and cleavage of both caspase-9 and caspase-3 due to disruption of mitochondrial membrane potential were significantly blocked by catalase and EUK-134 in BxPC-3 cells. Our results further demonstrate that capsaicin treatment not only inhibit the enzymatic activity and expression of SOD, catalase and glutathione peroxidase but also reduce glutathione level. Over-expression of catalase by transient transfection protected the cells from capsaicin-mediated ROS generation and apoptosis. Furthermore, tumors from mice orally fed with 2.5 mg/kg capsaicin show decreased SOD activity and an increase in GSSG/GSH levels as compared to controls. Taken together, our results suggest the involvement of mitochondrial complex-I and III in capsaicin-mediated ROS generation and decrease in antioxidant levels resulting in severe mitochondrial damage leading to apoptosis in pancreatic cancer cells

Molecular analysis of beta-globin gene mutations among Thai beta-thalassemia children: results from a single center study

Boonchai Boonyawat,1 Chalinee Monsereenusorn,2 Chanchai Traivaree2 1Division of Genetics, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand; 2Division of Hematology/Oncology, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand Background: Beta-thalassemia is one of the most common genetic disorders in Thailand. Clinical phenotype ranges from silent carrier to clinically manifested conditions including severe beta-thalassemia major and mild beta-thalassemia intermedia. Objective: This study aimed to characterize the spectrum of beta-globin gene mutations in pediatric patients who were followed-up in Phramongkutklao Hospital. Patients and methods: Eighty unrelated beta-thalassemia patients were enrolled in this study including 57 with beta-thalassemia/hemoglobin E, eight with homozygous beta-thalassemia, and 15 with heterozygous beta-thalassemia. Mutation analysis was performed by multiplex amplification refractory mutation system (M-ARMS), direct DNA sequencing of beta-globin gene, and gap polymerase chain reaction for 3.4 kb deletion detection, respectively. Results: A total of 13 different beta-thalassemia mutations were identified among 88 alleles. The most common mutation was codon 41/42 (-TCTT) (37.5%), followed by codon 17 (A>T) (26.1%), IVS-I-5 (G>C) (8%), IVS-II-654 (C>T) (6.8%), IVS-I-1 (G>T) (4.5%), and codon 71/72 (+A) (2.3%), and all these six common mutations (85.2%) were detected by M-ARMS. Six uncommon mutations (10.2%) were identified by DNA sequencing including 4.5% for codon 35 (C>A) and 1.1% initiation codon mutation (ATG>AGG), codon 15 (G>A), codon 19 (A>G), codon 27/28 (+C), and codon 123/124/125 (-ACCCCACC), respectively. The 3.4 kb deletion was detected at 4.5%. The most common genotype of beta-thalassemia major patients was codon 41/42 (-TCTT)/codon 26 (G>A) or betaE accounting for 40%. Conclusion: All of the beta-thalassemia alleles have been characterized by a combination of techniques including M-ARMS, DNA sequencing, and gap polymerase chain reaction for 3.4 kb deletion detection. Thirteen mutations account for 100% of the beta-thalassemia genes among the pediatric patients in our study. Keywords: mutation analysis, beta-globin gene, Thai childre

The effect of intravenous hydration strategy on plasma methotrexate clearance during intravenous high-dose methotrexate administration in pediatric oncology patients

Chanchai Traivaree,1 Napakjira Likasitthananon,2 Chalinee Monsereenusorn,1 Piya Rujkijyanont1 1Division of Hematology/Oncology, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand; 2Department of Pediatrics, Lat Krabang Hospital, Bangkok, Thailand Background: High-dose methotrexate (HD-MTX) is widely used as a standard chemotherapeutic agent in pediatric cancers. Most research studies have confirmed the therapeutic efficacy of HD-MTX; however, strategies to prevent side effects vary among institutions, especially in developing countries, with limited monitoring of plasma methotrexate level.Objective: To evaluate the effect of intravenous hydration during HD-MTX administration on plasma methotrexate clearance in pediatric oncology patients.Materials and methods: This study retrospectively reviewed 165 courses of HD-MTX administered to children with acute lymphoblastic leukemia (ALL), non-Hodgkin’s lymphoma (NHL), or osteosarcoma. Demographic data of patients were collected. Adverse complications related to HD-MTX and 72-hour plasma methotrexate level were analyzed between patients receiving intravenous hydration ≥3,000 mL/m2/day and those receiving hydration <3,000 mL/m2/day.Results: Among 56 HD-MTX (1.5 g/m2) courses in ALL, delayed methotrexate clearance was only found in one course administered with hydration <3,000 mL/m2/day. However, no correlation was observed between adverse complications and methotrexate levels. Of 34 HD-MTX (1.5–3 g/m2) courses in NHL, no significant correlation was observed between methotrexate levels and intravenous hydration. However, increased adverse complications were found in the course with delayed methotrexate clearance. Interestingly, among 75 HD-MTX (10–12 g/m2) courses in osteosarcoma, normal methotrexate clearance was successfully achieved in all courses administered with hydration ≥3,000 mL/m2/day compared with those administered with hydration <3,000 mL/m2/day (P=0.007). Furthermore, the courses administered with hydration <3,000 mL/m2/day and had delayed methotrexate clearance were more likely to develop adverse complications.Conclusion: Intravenous hydration of ≥3,000 mL/m2/day during HD-MTX administration is essentially required in osteosarcoma and can be considered as optional in ALL with HD-MTX <1.5 g/m2, especially in developing countries with limited monitoring of plasma methotrexate level. Keywords: high-dose methotrexate, hydration, children, cancer, osteosarcoma, leukemia, lymphom

Efficacy of Oral Acetaminophen and Intravenous Chlorpheniramine Maleate versus Placebo to Prevent Red Cell Transfusion Reactions in Children and Adolescent with Thalassemia: A Prospective, Randomized, Double-Blind Controlled Trial

Background. Thalassemia is a common congenital hemolytic disorder. In severe cases, regular blood transfusion is essentially required. The role of premedications to prevent transfusion reactions is varied among institutions with no standard guideline. Objective. To prospectively compare the risk of transfusion reactions in thalassemia patients premedicated with acetaminophen and chlorpheniramine maleate (CPM) versus placebo prior to blood transfusion. Material and Method. A randomized, double-blinded, placebo-controlled transfusion reaction study of 147 eligible patients was analyzed. All administered red blood cell (RBC) products were leukoreduced blood products. Patients were monitored and followed for the development of transfusion reactions for 24 hours after RBC transfusion. Results. A total of 73 patients randomized to receive active drugs consisting of acetaminophen and CPM were compared to 74 patients receiving placebo. The overall incidences of febrile reaction and urticarial rash were 6.9% and 22% in the patients randomized to receive active drugs comparing with 9.5% and 35.2% in the patients receiving placebo with no significant differences between two groups. However, delayed development of urticarial rash at 4-24 hours after RBC transfusion was significantly higher in female and patients receiving placebo. Conclusion. Administration of premedications in thalassemia patients receiving RBC transfusion without a history of transfusion reactions does not decrease the overall risk of transfusion reactions. However, the use of CPM might be beneficial to prevent delayed urticarial rash in those patients especially in females (Thai Clinical Trial Registry (TCTR) study ID: 20140526001)

Cholangiocarcinoma in a Child with Progressive Abdominal Distension and Secondary Hypercalcemia

Cholangiocarcinoma is extremely rare in childhood and has been reported in association with other underlying diseases. The survival and prognosis are dismal especially in patients with unresectable or advanced stage cholangiocarcinoma. Overall survival in patients with metastatic cholangiocarcinoma could be increased by using combination chemotherapy with cisplatin and gemcitabine. A case of childhood cholangiocarcinoma was hereby reported

Clinical and molecular genetic features of Hb H and AE Bart’s diseases in central Thai children

Chanchai Traivaree,1,* Boonchai Boonyawat,2,* Chalinee Monsereenusorn,1 Piya Rujkijyanont,1 Apichat Photia1 1Division of Hematology/Oncology, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand; 2Division of Genetics, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand *These authors contributed equally to this work Background: α-Thalassemia, one of the major thalassemia types in Thailand, is caused by either deletion or non-deletional mutation of one or both α-globin genes. Inactivation of three α-globin genes causes hemoglobin H (Hb H) disease, and the combination of Hb H disease with heterozygous hemoglobin E (Hb E) results in AE Bart’s disease. Objective: This study aimed to characterize the clinical and hematological manifestations of 76 pediatric patients with Hb H and AE Bart’s diseases treated at Phramongkutklao Hospital, a tertiary care center for thalassemia patients in central Thailand.Patients and methods: Seventy-six unrelated pediatric patients, 58 patients with Hb H disease and 18 patients with AE Bart’s disease, were enrolled in this study. Their clinical presentations, transfusion requirement, laboratory findings, and mutation analysis were retrospectively reviewed and analyzed.Results: A total of 76 pediatric patients with Hb H and AE Bart’s diseases who mainly lived in central Thailand were included in this study. The clinical severities of patients with non-deletional mutations were more severe than those with deletional mutations. Eighty-six percent of patients with non-deletional AE Bart’s disease required more blood transfusion compared to 12.5% of patients with deletional AE Bart’s disease. Non-deletional AE Bart’s disease also had a history of urgent blood transfusion with the average of 6±0.9 times compared to 1±0.3 times in patients with deletional Hb H disease. The difference was statistically significant.Conclusion: This study revealed the differences in clinical spectrum between patients with Hb H disease and those with AE Bart’s disease in central Thailand. The differentiation of α-thalassemia is essential for appropriate management of patients. The molecular diagnosis is useful for diagnostic confirmation and genotype–phenotype correlation. Keywords: genotype, phenotype, Hb H disease, AE Bart’s disease, Thai childre

Genotype–phenotype correlation among beta-thalassemia and beta-thalassemia/HbE disease in Thai children: predictable clinical spectrum using genotypic analysis

Chanchai Traivaree,1 Chalinee Monsereenusorn,1 Piya Rujkijyanont,1 Warakorn Prasertsin,2 Boonchai Boonyawat3 1Division of Hematology/Oncology, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand; 2Department of Pediatrics, Queen Savang Vadhana Memorial Hospital, Chonburi, Thailand; 3Division of Genetics, Department of Pediatrics, Phramongkutklao Hospital and College of Medicine, Bangkok, Thailand Introduction: Beta-thalassemia is a group of inherited hemolytic anemias and one of the most common genetic disorders in Thailand. The clinical spectrum of beta-thalassemia disease ranges from mild to severe clinical symptoms including mild beta-thalassemia intermedia (TI) and severe beta-thalassemia major (TM). Objective: This study aimed to determine the correlation between beta-globin gene (HBB) mutations and their phenotypic manifestations by evaluating patients’ clinical characteristics, transfusion requirements, growth and hematologic parameters, and hemoglobin typing among pediatric patients treated at Phramongkutklao Hospital. Materials and methods: Seventy beta-thalassemia patients, including 63 with beta-thalassemia/hemoglobin E (HbE) and 7 with either homozygous or compound heterozygous beta-thalassemia, were enrolled in this study. Their clinical presentation, growth parameters and laboratory findings were reviewed and analyzed. The mean follow-up time was 10.52±5.62 years. Mutation analysis in each individual was performed using multiplex amplification refractory mutation system (M-ARMS), direct DNA sequencing of beta-globin gene and gap PCR for 3.4 kb deletion detection. Results: All 7 homozygous and compound heterozygous beta-thalassemia patients were classified in TM. Among 63 patients with beta-thalassemia/HbE, 58 were classified in TM and 4 were classified in TI. Mean age at diagnosis was 0.8±0.49 years for homozygous or compound heterozygous beta-thalassemia and 3.43±3.5 years for beta-thalassemia/HbE. The most common HBB mutation was HBB:c.126_129delCTTT [codon 41/42 (-TCTT)] found in 34 alleles (48.6%). The height for age was also lower in homozygous beta-thalassemia patients (<3rd percentile) compared to compound heterozygous beta-thalassemia patients (25–50th percentile). Conclusion: This study revealed a genotype–phenotype correlation of the most prevalent beta-thalassemia in Thai children using diagnostic capacity in genotypic analysis of HBB mutation. Our findings can provide a better prediction of clinical manifestation and severity by early identification of the type of the HBB mutations. Keywords: beta-thalassemia disease, hemoglobinopathy, mutation, clinical, childre