Serum soluble transferrin receptor concentration as a biomarker of erythropoietic activity: surrogate marker of adequate transfusion in adult Beta-thalassaemia intermedia patients

Management of Beta (β)-thalassaemia intermedia in contrast to β-thalassaemia major patients has no clear guidelines as to indicators of adequate transfusion. Regular blood transfusion suppresses bone marrow erythropoietic activity. Serum soluble transferrin receptor (sTfR) concentration is a marker for erythropoietic activity, with increased sTfR being associated with functional iron deficiency and increased erythropoietic activity. This study aimed to determine the use of sTfR as an indicator of adequate transfusion in adult β-thalassaemia intermedia patients. A cross-sectional study was conducted at Hospital Ampang, Malaysia, for six months. Patient group included six β-thalassaemia intermedia and 34 HbE-β-thalassaemia transfused patients. None of the patients were on regular monthly blood transfusions as in β-thalassaemia major. The control group comprised of 16 healthy subjects with normal haematological parameters. Haemoglobin (Hb) analysis, sTfR and ferritin assays were performed. Hb and HbA percentages (%) were found to be significantly lower in patients compared to the controls, while HbE%, HbF%, sTfR and ferritin were significantly higher in patients. An inverse relationship was found in the controls between HbF% with Hb (r = -0.515, p < 0.05) and HbA% (r = -0.534, p < 0.05). In patients, sTfR showed an inverse relationship with HbA% (r = -0.618, p = 0.000) and a positive correlation with HbE% (r = 0.418, p = 0.007) and HbF% (r = 0.469, p = 0.002). Multivariate analysis showed that HbA% (r = 2.875, p = 0.048), HbE% (r = 2.872, p = 0.020) and HbF% (r = 2.436, p = 0.013) best predicted sTfR independently in patients. Thus, sTfR is a useful marker for erythropoiesis. The elevate sTfR in these patients indicate that the transfusion regimen used was inadequate to suppress ineffective erythropoiesis. Hb levels may not be the best target for monitoring transfusion treatment in β-thalassaemia intermedia patients, but the use of sTfR is helpful in individualising transfusion regimens

Case series of homozygous and compound heterozygosity of Hb Malay, the diagnostic features and transfusion requirements

Hb Malay was first described in 1989 following an investigation of anaemia in a 22-year-old Malay gentleman who was homozygous for this b chain variant. This Hb variant is caused by AAC à AGC mutation at codon 19 of the b globin gene resulting in the substitution of serine for asparagine [1]. The mutation creates cryptic RNA splice site in exon 1 of the b-globin gene leading to an abnormal RNA processing. Thus, this mutation not only produces variant haemoglobin but also a mild b+ thalassemia phenotype [2]. A retrospective analysis was carried out at the Clinical Haematology Laboratory, Hospital Ampang, Selangor, Malaysia from 2012 to 2015. A total of 12 cases of confirmed heterozygous, homozygous or compound heterozygous of Hb Malay were collected. The diagnostic workups in this centre included complete blood count (CBC), blood smear, haemoglobin analysis and molecular study. Table 1 shows the haematological characteristics of Hb Malay and its combination with other thalassaemia/ haemoglobinopathies. Table 2 shows the clinical features and transfusion requirements of various presentation of Hb Malay. The definitive diagnosis of Hb Malay can only be made by molecular analysis. Both reverse phase high performance liquid chromatography (HPLC) for haemoglobin variant and capillary zone electrophoresis (CZE) cannot differentiate between Hb A and Hb Malay as it is co-migrated. Previously, it was reported that there was an increased production of Hb F between 12-32% in cases of homozygous Hb Malay and compound heterozygous Hb E/Malay [3]. In our case, the Hb F levels in homozygous Hb Malay were 47.4% and 30%, respectively (Case 1 and 2). Patients with homozygous Hb Malay were non-transfusion dependent with average haemoglobin of 7 to 8g/dL, whereas in Hb Malay trait the haemoglobin level was normal. The Hb E/ Malay patients were also asymptomatic, although the average haemoglobin was lower (10g/dL) compared to the classical Hb E trait (12.4g/dL) [5]. Hb F level in Hb E/ Malay was reported to be above 12% [3], in one of our cases, the Hb F was only 3.3% (Case 4). Molecular analysis showed this patient was also homozygous for 158 Gγ Xmn polymorphism, as well as -α3.7 deletion. This was rather an interesting finding as this polymorphism was associated with higher level of Hb F [4]. Similarly, the presence of alpha thalassaemia was reported to reduce the Hb E percentage to less than 25%, but in this case the Hb E was 52.6% [5]. Compound heterozygous of Hb Malay/ Beta thalassaemia resulted in thalassaemia intermedia with variable phenotypes depending on the type of mutations. The other 2 combinations of Hb Malay with Hb S and HPFH also presented as thalassaemia intermedia. These findings were in keeping with the phenotype of Hb Malay resembling b+ thalassaemia

Correlation of BACH1 and hemoglobin E/Beta-thalassemia globin expression

Objective: The diverse clinical phenotype of hemoglobin E (HbE)/β-thalassemia has not only confounded clinicians in matters of patient management but has also led scientists to investigate the complex mechanisms involved in maintaining the delicate red cell environment where, even with apparent similarities of α- and β-globin genotypes, the phenotype tells a different story. The BTB and CNC homology 1 (BACH1) protein is known to regulate α- and β-globin gene transcriptions during the terminal differentiation of erythroid cells. With the mutations involved in HbE/β-thalassemia disorder, we studied the role of BACH1 in compensating for the globin chain imbalance, albeit for fine-tuning purposes. Materials and Methods: A total of 47 HbE/β-thalassemia samples were analyzed using real-time quantitative polymerase chain reaction and correlated with age, sex, red blood cell parameters, globin gene expressions, and some clinical data. Results: The BACH1 expression among the β-thalassemia intermedia patients varied by up to 2-log differences and was positively correlated to age; α-, β-, and γ-globin gene expression level; and heme oxygenase 1 protein. BACH1 was also negatively correlated to reticulocyte level and had a significant correlation with splenectomy. Conclusion: This study indicates that the expression of BACH1 could be elevated as a compensatory mechanism to decrease the globin chain imbalance as well as to reduce the oxidative stress found in HbE/β-thalassemia

Human hemoglobin G-Makassar variant masquerading as sickle cell anemia

Human hemoglobin of G-Makassar variant has been reported very rarely with Beta Thalassemia. In year 1969 Hb GMakassar was first identified in Makassar, Sulawesi (Celebes), Republic of Indonesia. The disease was first published in 1969 and 33 years later it has been reported at a family of Thailand origin. We report a 45-yearold Malay man who was investigated for anemia and thrombocytopenia then diagnosed with Hb G-Makassar. This finding describes as a new Hemoglobin GMakassar discovered in Malaysia after 14 years diagnosed in Thailand

Effects of transfusion and splenectomy on globin chain expression in NTDT HbE/β-thalassaemia

Introduction: Majority of HbE/β-thalassaemia patients resembles the phenotype of non-transfusion dependent thalassaemia (NTDT). Current management strategies are highly diverse, and the objective of this study is to examine the effects of different treatments on multiple parameters in NTDT HbE/β-thalassaemia to further streamline the management of this disorder. Methods: In this cross-sectional study, we analysed the correlation between different treatment strategies with variable parameters including haematological parameter and globin gene expression. Statistical analyses were carried out using non-parametric tests such as Kruskal-Wallis and Mann-Whitney tests. Results: A total of 29 HbE/β-thalassaemia patients were included in the study. Data showed statistically significant differences were observed in the MCV, MCHC levels, reticulocyte count and log α/β fold change between the groups. Further analysis showed higher log α/β fold change in the transfusion only group compared to the non-treated group. Red blood cell count was found to be lower in transfused and splenectomised group compared to transfusion only. Significantly higher MCV level and reticulocyte count was seen in transfusion and splenectomised group compared to both non-treated and transfusion only groups and higher MCH level in the transfusion and splenectomised group compared to transfusion only group. Conclusion: In general, regardless of single or double combined therapies, HbE/β-thalassaemia showed variable changes in laboratory parameters to the therapies received particularly splenectomy

Genomic Alterations, Gene Expression Profiles and Functional Enrichment of Normal-Karyotype Acute Myeloid Leukaemia Based on Targeted Next-Generation Sequencing

Characterising genomic variants is paramount in understanding the pathogenesis and heterogeneity of normal-karyotype acute myeloid leukaemia (AML-NK). In this study, clinically significant genomic biomarkers were ascertained using targeted DNA sequencing and RNA sequencing on eight AML-NK patients’ samples collected at disease presentation and after complete remission. In silico and Sanger sequencing validations were performed to validate variants of interest, and they were followed by the performance of functional and pathway enrichment analyses for overrepresentation analysis of genes with somatic variants. Somatic variants involving 26 genes were identified and classified as follows: 18/42 (42.9%) as pathogenic, 4/42 (9.5%) as likely pathogenic, 4/42 (9.5%) as variants of unknown significance, 7/42 (16.7%) as likely benign and 9/42 (21.4%) as benign. Nine novel somatic variants were discovered, of which three were likely pathogenic, in the CEBPA gene with significant association with its upregulation. Transcription misregulation in cancer tops the affected pathways involving upstream genes (CEBPA and RUNX1) that were deregulated in most patients during disease presentation and were closely related to the most enriched molecular function gene ontology category, DNA-binding transcription activator activity RNA polymerase II-specific (GO:0001228). In summary, this study elucidated putative variants and their gene expression profiles along with functional and pathway enrichment in AML-NK patients.This research was funded by Universiti Sains Malaysia, grant number GIPS-PhD 311/PPSP/4404814 (23 April 2020). Universiti Sains Malaysia funded the APC

Turoctocog alfa pegol provides effective management for major and minor surgical procedures in patients across all age groups with severe haemophilia A:Full data set from the pathfinder 3 and 5 phase III trials

Introduction Turoctocog alfa pegol is a glycoPEGylated recombinant factor VIII (FVIII) with an extended half-life developed for prophylaxis, treatment of bleeds and perioperative management in patients with haemophilia A. Aim Evaluate the efficacy and safety of turoctocog alfa pegol treatment for major and minor surgeries in the pathfinder 3 and 5 phase III trials. Methods Adults/adolescents aged >= 12 years with severe haemophilia A (FVIII 80% during major surgery (pathfinder 3). The primary end point was haemostatic efficacy during surgery; secondary end points were blood loss, haemostatic effect postsurgery, consumption, transfusions, safety and health economics. Children (0-11 years) undergoing minor surgeries received 20-75 IU/kg turoctocog alfa pegol at Investigator's discretion (pathfinder 5). Results pathfinder 3 included 35 patients undergoing 49 major surgeries. Haemostasis was successful in 47/49 (95.9%) surgeries; two had moderate haemostatic responses. Median (mean) blood loss during major surgery was 75 (322.6) mL. Four bleeds were reported postsurgery; three were successfully treated with turoctocog alfa pegol (one was not evaluated). On the day of surgery, overall mean (median) dose was 75.5 (74.5) IU/kg and mean (median) number of doses was 1.7 (2.0). Five procedures required 11 transfusions on the day of surgery or days 1-6. No safety concerns or inhibitors were identified. Forty-five minor surgeries in 23 children were performed without complications. Conclusion Turoctocog alfa pegol was effective for perioperative haemostatic management of major and minor surgeries in patients across age groups with severe haemophilia A

Microarray-based genomic analysis identifies germline and somatic copy number variants and loss of heterozygosity in acute myeloid leukaemia

Introduction: Insights into molecular karyotyping using comparative genomic hybridization (CGH) and single nucleotide polymorphism (SNP) arrays enable the identification of copy number variations (CNVs) at a higher resolution and facilitate the detection of copy neutral loss of heterozygosity (CN-LOH) otherwise undetectable by conventional cytogenetics. The applicability of a customised CGH+SNP 180K DNA microarray in the diagnostic evaluation of Acute Myeloid Leukaemia (AML) in comparison with conventional karyotyping was assessed in this study. Methods: Paired tumour and germline post induction (remission sample obtained from the same patient after induction) DNA were used to delineate germline variants in 41 AML samples and compared with the karyotype findings. Results: After comparing the tumour versus germline DNA, a total of 55 imbalances (n 5-10 MB = 21, n 10-20 MB = 8 and n >20 MB = 26) were identified. Gains were most common in chromosome 4 (26.7%) whereas losses were most frequent in chromosome 7 (28.6%) and X (25.0%). CN-LOH was mostly seen in chromosome 4 (75.0%). Comparison between array CGH+SNP and karyotyping revealed 20 cases were in excellent agreement and 13 cases did not concord whereas in 15 cases finding could not be confirmed as no karyotypes available. Conclusion: The use of a combined array CGH+SNP in this study enabled the detection of somatic and germline CNVs and CN-LOHs in AML. Array CGH+SNP accurately determined chromosomal breakpoints compared to conventional cytogenetics in relation to presence of CNVs and CN-LOHs

Recommendations from the International Consensus Conference on Anemia Management in Surgical Patients (ICCAMS)

Background: Perioperative anemia has been associated with increased risk of red blood cell transfusion and increased morbidity and mortality following surgery. The optimal approach to the diagnosis and management of perioperative anemia is not fully established. Objective: To develop consensus recommendations for anemia management in surgical patients. Methods: An international expert panel reviewed the current evidence and developed recommendations using modified RAND Delphi methodology. Results: The panel recommends that all patients be screened for anemia prior to surgery. Appropriate therapy for anemia should be guided by an accurate diagnosis of the etiology. The need to proceed with surgery in some patients with anemia is expected to persist. However, early identification and effective treatment of anemia has the potential to reduce the risks associated with surgery and improve clinical outcomes. As with preoperative anemia, postoperative anemia should be treated in the perioperative period. Conclusions: Early identification and effective treatment of anemia has the potential to improve clinical outcomes in surgical patients

Clinical evaluation of dengue and identification of risk factors for severe disease: protocol for a multicentre study in 8 countries

Background: The burden of dengue continues to increase globally, with an estimated 100 million clinically apparent infections occurring each year. Although most dengue infections are asymptomatic, patients can present with a wide spectrum of clinical symptoms ranging from mild febrile illness through to severe manifestations of bleeding, organ impairment, and hypovolaemic shock due to a systemic vascular leak syndrome. Clinical diagnosis of dengue and identification of which patients are likely to develop severe disease remain challenging. This study aims to improve diagnosis and clinical management through approaches designed a) to differentiate between dengue and other common febrile illness within 72 h of fever onset, and b) among patients with dengue to identify markers that are predictive of the likelihood of evolving to a more severe disease course. Method/Design: This is a prospective multi-centre observational study aiming to enrol 7–8000 participants aged ≥ 5 years presenting with a febrile illness consistent with dengue to outpatient health facilities in 8 countries across Asia and Latin America. Patients presenting within 72 h of fever onset who do not exhibit signs of severe disease are eligible for the study. A broad range of clinical and laboratory parameters are assessed daily for up to 6 days during the acute illness, and also at a follow up visit 1 week later. Discussion: Data from this large cohort of patients, enrolled early with undifferentiated fever, will be used to develop a practical diagnostic algorithm and a robust clinical case definition for dengue. Additionally, among patients with confirmed dengue we aim to identify simple clinical and laboratory parameters associated with progression to a more severe disease course. We will also investigate early virological and serological correlates of severe disease, and examine genetic associations in this large heterogeneous cohort. In addition the results will be used to assess the new World Health Organization classification scheme for dengue in practice, and to update the guidelines for “Integrated Management of Childhood Illness” used in dengue-endemic countries. Trial registration: NCT01550016. Registration Date: March 7, 201