PUM1 Mediates the Posttranscriptional Regulation of Human Fetal Hemoglobin

The fetal-to-adult hemoglobin switching at about the time of birth involves a shift in expression from γ-globin to β-globin in erythroid cells. Effective re-expression of fetal γ-globin can ameliorate sickle cell anemia and β-thalassemia. Despite the physiological and clinical relevance of this switch, its posttranscriptional regulation is poorly understood. Here, we identify Pumilo 1 (PUM1), an RNA-binding protein with no previously reported functions in erythropoiesis, as a direct posttranscriptional regulator of β-globin switching. PUM1, whose expression is regulated by the erythroid master transcription factor erythroid Krüppel-like factor (EKLF/KLF1), peaks during erythroid differentiation, binds γ-globin messenger RNA (mRNA), and reduces γ-globin (HBG1) mRNA stability and translational efficiency, which culminates in reduced γ-globin protein levels. Knockdown of PUM1 leads to a robust increase in fetal hemoglobin (∼22% HbF) without affecting β-globin levels in human erythroid cells. Importantly, targeting PUM1 does not limit the progression of erythropoiesis, which provides a potentially safe and effective treatment strategy for sickle cell anemia and β-thalassemia. In support of this idea, we report elevated levels of HbF in the absence of anemia in an individual with a novel heterozygous PUM1 mutation in the RNA-binding domain (p.(His1090Profs∗16); c.3267_3270delTCAC), which suggests that PUM1-mediated posttranscriptional regulation is a critical player during human hemoglobin switching

Multispectral Imaging for Microchip Electrophoresis Enables Point-of-Care Newborn Hemoglobin Variant Screening

Hemoglobin (Hb) disorders affect nearly 7% of the world\u27s population. Globally, around 400,000 babies are born annually with sickle cell disease (SCD), primarily in sub-Saharan Africa where morbidity and mortality rates are high. Screening, early diagnosis, and monitoring are not widely accessible due to technical challenges and cost. We hypothesized that multispectral imaging will allow sensitive hemoglobin variant identification in existing affordable paper-based Hb electrophoresis. To test this hypothesis, we developed the first integrated point-of-care multispectral Hb variant test: Gazelle-Multispectral. Here, we evaluated the accuracy of Gazelle-Multispectral for Hb variant newborn screening in 265 newborns with known hemoglobin variants including hemoglobin A (Hb A), hemoglobin F (Hb F), hemoglobin S (Hb S) and hemoglobin C (Hb C). Gazelle-Multispectral detected levels of Hb A, Hb F, Hb S, and Hb C/E/A2, demonstrated high correlations with the results reported by laboratory gold standard high performance liquid chromatography (HPLC) at Pearson Correlation Coefficient = 0.97, 0.97, 0.93, and 0.95. Gazelle-Multispectral demonstrated accuracy of 96.8% in subjects of 0–3 days, and 96.9% in newborns. The ability to obtain accurate results on newborn samples suggest that Gazelle-Multispectral can be suitable for large-scale newborn screening and for diagnosis of SCD in low resource settings

Antithrombin-III Mitigates Thrombin-Mediated Endothelial Cell Contraction and Sickle Red Blood Cell Adhesion in Microscale Flow

Individuals with sickle cell disease (SCD) have persistently elevated thrombin generation that results in a state of systemic hypercoagulability. Antithrombin-III (ATIII), an endogenous serine protease inhibitor, inhibits several enzymes in the coagulation cascade, including thrombin. Here, we utilize a biomimetic microfluidic device to model the morphology and adhesive properties of endothelial cells (ECs) activated by thrombin and examine the efficacy of ATIII in mitigating the adhesion of SCD patient-derived red blood cells (RBCs) and EC retraction. Microfluidic devices were fabricated, seeded with ECs, and incubated under physiological shear stress. Cells were then activated with thrombin with or without an ATIII pretreatment. Blood samples from subjects with normal haemoglobin (HbAA) and subjects with homozygous SCD (HbSS) were used to examine RBC adhesion to ECs. Endothelial cell surface adhesion molecule expression and confluency in response to thrombin and ATIII treatments were also evaluated. We found that ATIII pretreatment of ECs reduced HbSS RBC adhesion to thrombin-activated endothelium. Furthermore, ATIII mitigated cellular contraction and reduced surface expression of von Willebrand factor and vascular cell adhesion molecule-1 (VCAM-1) mediated by thrombin. Our findings suggest that, by attenuating thrombin-mediated EC damage and RBC adhesion to endothelium, ATIII may alleviate the thromboinflammatory manifestations of SCD

Prevalence of relative systemic hypertension in adults with sickle cell disease in Ghana - Fig 1

<p><b>Trends in probability of having normal BP with advancing age (A) and stratification of study population by blood pressure categories and age (B)</b>. Relative hypertension is denoted in (A) as borderline hypertension. Blood Pressure values were adjusted for hemoglobin and WBC count.</p

Study population characteristics.

<p>Study population characteristics.</p

Percentage of different complications by blood pressure category.

<p>Percentage of different complications by blood pressure category.</p

Mean pulse pressure distribution by gender and age, adjusted for hemoglobin.

<p>The median (IQR) for pulse pressure was 45 mmHg (39–55) and in linear regression analysis pulse pressure was associated with total hemoglobin (beta = -0.4 per mg/dl, P = 0.031), age (beta = 0.3 per year, P < 0.001) and male gender (beta = 4.4, P < 0.001) (Fig 2).</p

DataSheet_1_Case report: Daratumumab treatment in pre-transplant alloimmunization and severe hemolytic anemia.pdf

Daratumumab, a CD38 monoclonal antibody that has been FDA-approved to treat multiple myeloma, has acquired popularity and is used off-label for both auto- and alloantibody mediated disorders, particularly in refractory/resistant circumstances. Much of the published data for its use in pediatric blood disorders has been in post-transplant autoimmune cytopenias. Here we describe three patients in whom daratumumab was used outside of post-transplant autoimmune cytopenias, highlighting further potential uses of this medication.</p

Grndad and Disease Modifying Therapy (DMT): Shifts in Dmt Are Seen at the Adolescent/Young Adult Transition in Sickle Cell Disease in a Multi-Site Prospective Registry

The ability to characterize the modern person living with SCD in the US has been limited by the lack of a well-curated longitudinal registry. The Globin Research Network for Data and Discovery ( GRNDaD) registry aims to overcome this challenge by collecting data from the now 53 IRB-approved centers across the US in collaboration with the National Alliance of Sickle Cell Centers (NASCC) and the HRSA-funded Sickle Cell Disease Treatment Demonstration Project Grant. Here, we describe the use of disease-modifying therapy in (actively consented) adults and children with Hgb SS/ SB 0 thalassemia (SCA) from 37 sites. Methods: Each site consents subjects and enters extensive baseline data including SCD complications and labs, subjects also complete patient-reported outcomes (PROs)-both at baseline and annually. Each subject is expected to have an annual follow-up where data is extracted from the Electronic Health Record (EHR). All data is stored in REDCap and, for this abstract, data were extracted into R on all active subjects (data entry in the last two years) who have complete minimum data collected. Median values were compared using Mann-Whitney U tests. Disease-modifying therapy (DMT) in this study comprises hydroxyurea (HU), chronic red blood cell (RBC) transfusions, crizanlizumab, L-glutamine, and voxelotor. Results: There are 2501 active patients in GRNDaD. Twenty-six percent of subjects are pediatric </= 18 years of age, 55.5% are female and 66.7% have HgbSS or HgbSB 0 thalassemia (SCA). Here, we report data on people with SCA only, whose minimum data were complete (n=1272). Table 1 shows DMT by age. 187 (21%) adults and 31 (8.5%) children were not on DMT. Figure 2 is a box plot of the absolute neutrophil count (ANC) comparing adults and children who are and are not taking HU. Adults on HU had significantly lower ANCs than those not on HU (median = 4.4 (IQR: 3.07, 6.3) v 6.6 (IQR:4.07, 8.14), p<0.001) (Figure). Similar findings were observed for children ( median = 3.5 (IQR:2.5, 5.7) Vs 8.1 (5.8, 12.8) , p =0.04) Both children and adults on HU had higher MCVs than those not on HU (median for peds: 91.5 (IQR: 85.1, 98.75) v 86 (IQR: 83.3, 86.1), p=0.05, median for adults: 96.5 (IQR: 89, 107) v 90 (IQR: 86.1, 95.1), p<0.001). We compared hemoglobin for those on and off HU, and on and off voxelotor, excluding patients on chronic transfusion therapy. There was a statistically significant difference in hemoglobin when comparing on or off of HU (median 8.9 g/dl (IQR 7.95, 9.9) v 8.2 g/dl (IQR: 7.2, 9.7, p =0.047)). There were no statistically significant differences in hemoglobin when comparing subjects on or off of voxelotor (median = 8.4 g/dl (IQR: 7.5, 9.6) vs 7.9 (IQR: 7.4, 9.8), p =0.57). In the adults, there were statistically significantly more males than females on HU, but there were no differences seen in the pediatric cohort. Examining DMT by age group, chronic RBC transfusion use doubled from the 11-17 age group to the 18-29-year age group (12% to 25%) and HU use decreased (83%-59%) over this same period. Limitations: This cohort may over-represent those on DMT as those enrolled in GRNDaD were more likely to have clinic visits. Discussion: GRNDaD has doubled the number of sites consenting subjects and entering data over the last year. The number is expected to increase again in the next 12 months, as an additional 15 sites are already IRB-approved with a goal to include all SCD centers in NASCC. This cross-sectional description of the current population of people living with SCD and treated in NASCC-recognized SCD centers in the US shows that the majority of those with SCA are receiving DMT. There is a noticeable decrease in HU use during the transition period with an increase in the use of chronic transfusion therapy. Newer DMT has had limited uptake in this cohort. The next steps will be to provide longitudinal data on this population and examine associations of DMT and important clinical outcomes including PROs