ALBERT

Description: Sickle Cell Anemia (SCA) is a leading cause of childhood stroke in sub-Saharan Africa and sickle cell brain vasculopathy manifests either as overt stroke or clinically "silent infarcts". This study aimed to describe brain abnormalities seen on magnetic resonance imaging in Ugandan SCA children. Our hypothesis was that multi-model abnormalities would be associated with cerebrovasculopathy found on MRI/MRA. Methods As part of a larger study to determine the burden and spectrum of neurological and cognitive impairments in SCA children in Uganda, we selected 81 children ages 1-12 years with HbSS, a sample enriched for possible brain pathology from Mulago hospital SCA clinic out of a random sample of 265 stable children. None was receiving hydroxyurea. All had detailed clinical history, and physical, neurological and cognitive testing, trans-cranial Doppler (TCD) cerebral blood flow velocity determination and non-contrasted brain MRI/MRA using a 1.5 Tesla scanner. Cognitive testing was performed using age-specific tools validated for Ugandan children. Results Of the 81 participants imaged, 61 had one or more of history of stroke, an abnormal neurological exam, cognitive impairment or abnormal TCD, while 20 had normal test results. MR abnormalities were seen in 35/61 (63.9%) participants with probable brain pathology and in 4/20 (20.0%) without any probable brain pathology. They included different structural abnormalities seen in all brain regions ranging from only T2 weighted hyper-intensities, white matter lacuna infarction to bilateral ischemic and multi-focal cerebral infarcts with associated compensatory hydrocephalus. MRA abnormalities ranged from cerebral microangiopathy to multiple stenosis and occlusions of major arteries, including moya-moya in 4 subjects. Severe vessel obstructions were also seen in multiple young children

Description: Introduction. Hydroxyurea treatment has proven safety, feasibility, and efficacy for children with sickle cell anemia living in sub-Saharan Africa. Even in malaria endemic regions, hydroxyurea is safe and has been shown to reduce vaso-occlusive events and lower mortality. The optimal dosing regimen is not known, however, and specifically whether a fixed dose at 15-20 mg/kg/day is better than escalation to maximum tolerated dose (MTD) at 25-30 mg/kg/day. Particularly in low-resource settings, additional drug-related toxicities and monitoring costs associated with dose escalation could outweigh potential laboratory and clinical benefits. Methods. Children with sickle cell anemia previously enrolled in NOHARM (NCT01976416) received hydroxyurea at ~20 mg/kg/day during the open-label portion of that trial, after which they transitioned to commercial supply at that same daily dose. All were then offered enrollment in the NOHARM MTD trial (NCT03128515), a double-blinded trial with 1:1 randomization between continuing fixed-dose oral hydroxyurea (20 ± 2.5 mg/kg/day) versus dose-escalation to MTD (30 ± 2.5 mg/kg/day), using hydroxyurea tablets donated by Addmedica. The primary study outcome was the proportion of study participants who achieved either hemoglobin ≥9.0 g/dL or fetal hemoglobin ≥20% after 24 months of treatment. Secondary outcomes included sickle-related clinical adverse events, malaria events, and laboratory toxicities. We now present the main results of the NOHARM MTD trial. Results. A total of 187 children enrolled and began study treatment: 94 children (4.8 ± 0.9 years, 55 males) received hydroxyurea at fixed dose versus 93 who received hydroxyurea with dose escalation (4.6 ± 1.0 years, 47 males). At Month 6 of study treatment the average doses were 19.3 ± 1.7 mg/kg/day and 29.0 ± 3.5 mg/kg/day, respectively, and these dose differences were continued through Month 12 and Month 18. The trial was terminated prematurely by the independent Data Safety Monitoring Board after 146 children had reached Month 18, when the clinical complications and interventions were significantly fewer among children randomized to MTD compared to fixed-dose. The incidence rate ratio (IRR) with 95% confidence intervals (CI) included the following: all sickle-related Adverse Events [IRR = 0.43, CI = (0.34, 0.56), P

Description: Background: Children with sickle cell anemia (SCA) are highly susceptible to stroke, impaired neurocognitive function and other manifestations of pediatric cerebral vasculopathy. Detailed multifaceted evaluations of children with SCA in sub-Saharan Africa are limited, including neurocognitive testing. Methods: We aimed to establish the prevalence and types of neurocognitive dysfunction in a cross-sectional study in a randomly selected sample of patients with HbSS ages 1-12 years at a large SCD clinic in Kampala, Uganda. Neurocognitive function was assessed by Mullen Scales of Early Learning (ages 1-4) and Kaufman Assessment Battery for Children, 2nd edition (ages 5-12) using linguistically validated local translations (John CC, Pediatrics 2008; Bangirana P., Clin Infect Dis. 2014). The Behavioral Rating Inventory for Executive Function was used to assess executive function. Children also underwent standardized stroke examination (PedsNIHSS) and transcranial Doppler ultrasound (TCD) for measuring stroke risk by arterial velocities in pediatric SCD. Results: Experienced testers performed neurocognitive assessment in 242 participants: 100 (41%) ages of 1-4 years and 142 ages 5-12 years (58.7%). Mean age was 5.44 ±2.9 years; 51.6% were male. Mean hemoglobin was 7.3 ±1.02 g/dl; 76.5% had hemoglobin

Description: Key Points Compared with placebo, hydroxyurea did not increase the incidence or severity of malaria events in Ugandan children with SCA. Hydroxyurea provided significant clinical and laboratory benefits, suggesting it will be safe and effective across sub-Saharan Africa.

Description: Introduction: Despite high prevalence of Sickle Cell Anemia (SCA) in sub-Saharan Africa, systematic evaluation of structural and functional impact of pediatric sickle cerebrovasculopathy has been limited. In a cross-sectional study of children with HbSS, ages 1-12 years, receiving care at the Mulago Hospital SCD clinic in Kampala, we hypothesized there would be high prevalence of multi-modal brain injury and stroke risk, as defined by 4 different adverse outcomes. Here we present our completed study results. Methods: Patients were randomly selected from the electronic clinic roster. None was taking hydroxyurea. Exclusion criteria: acute illness, Hb 2 Z scores below age-specific established community norms. Transcranial doppler ultrasounds (TCDs): Non-imaging TCDs (SonaraTek) were performed by two study trained staff (excellent inter-operator reliability 〉0.85); read by a U.S.-certified co-investigator. Brain magnetic resonance imaging (MRI), including arterial imaging (MRA): Subset of 81 (30.1%) of subjects (n=81), enriched for 1 or more testing abnormality, underwent non-contrast MRI/MRA imaging by a single 1.5T scanner. Two radiologists performed clinical reads. Statistical methods: ANOVA, multivariate, chi square, logistic regression. Results: Of 287 patients screened, all 265 eligible subjects were enrolled. Mean age was 5.5 SD 2.9 years, 52.3% male (Table 1). Mean Hb was 7.3 SD1.0 gm/dl. Each outcome was normally distributed.Stroke exam: Results: see below. Neurocognitive results: Overall results: see below. Poor testing: 2 of 100 ages 1-4 (2.0%) and 25 of 144 ages 5-12 (17.4%). TCDs: 13 of 251 (5.2%) were unread due to inadequate bone windows (12), poorly cooperative subject (1). Overall results: see below. MRI/MRAs: Overall results: see below. Abnormalities ranged from multiple small infarcts to multiple, bilateral infarcts and multi-vessel arterial stenoses. Among those with abnormal MRI/MRA, 13 (29.5%) also had a non-normal TCD. Summary data: In all, 58 (21.9%) subjects had ³1 abnormal outcomes. Among the 81 subjects with MR imaging, 28 (34.6%) had ³2 abnormal outcomes: 18 had 2; 9 had 3; 1 had all 4. Analyses: Abnormal neurocognitive results were highly dependent (6.8-fold) on age (

Description: Background: Children with sickle cell anemia (SCA) are highly susceptible to stroke and other manifestations of pediatric cerebral vasculopathy. Detailed evaluations of children with SCA in sub-Saharan Africa are limited, especially magnetic resonance imaging and angiography (MRI/MRA). In a sample of Ugandan children receiving care at the Mulago Hospital sickle cell clinic in Kampala and were not on disease-modifying therapy, we examined the range of MR imaging findings, and how those findings correlated with standardized demographic, clinical, neurological and neurocognitive assessments. Methods: From within a larger sample of 265 participants with HbSS ages 1-12 years not taking disease-modifying therapy and enrolled in the BRAIN SAFE study, a sub-sample of 81 underwent non-contrast MRI/MRA on a 1.5 Telsa scanner. Participants also underwent 3 standardized assessments: neurocognitive testing by experienced testers using the Mullen Scales of Early Learning (for ages 1-4 years) or Kaufman Assessment Battery for Children, 2nd edition (for ages 5-12) (abnormal z-score of -2 or lower), stroke examination (PedsNIHSS) and transcranial Doppler ultrasound (TCD) using criteria for pediatric SCA. Participants undergoing MRI/MRA intentionally included 29 without any abnormal findings. MRI scans included T1- and T2- weighted images, T2 FLAIR and MRA three-dimensional time-of-flight technique. MR scans were interpreted by clinical and research methods, the latter per SWiTCH protocol (Helton, Blood 2014). Adjudication of differing reads was performed by a blinded third neuroradiologist. Results: A total of 81 children with SCA were examined by MRI/MRA. Mean age was 6.48 ± 2.75 years; 50.6% were male. Mean hemoglobin was 7.26±0.90 g/dl; 75% had hemoglobin

Description: Background: Children with sickle cell anemia (SCA) are highly susceptible to stroke and other manifestations of pediatric cerebral vasculopathy. Inflammation is a risk factor for acute stroke. In a cross-sectional study of Ugandan children, we asked whether inflammatory markers correlated with findings on neurological and neurocognitive testing and magnetic resonance imaging (MRI). Methods: Stored plasma samples were obtained at a well study visit from 81 children, a subset within a larger sample of study participants with HbSS ages 1-12 years recruited at the Mulago Hospital sickle cell clinic in Kampala. Participants underwent 4 standardized assessments: neurocognitive testing using the Mullen Scales of Early Learning (for ages 1-4 years) or Kaufman Assessment Battery for Children, 2nd edition (for ages 5-12) (abnormal at z-score of -2 or lower), examination for prior stroke (PedsNIHSS) and transcranial Doppler ultrasound (TCD) using criteria for pediatric SCA, plus a non-contrast MRI/MRA on a 1.5 Telsa scanner, analyzed for T1- and T2- weighted images and T2 FLAIR. MR scans were interpreted by clinical and research methods, the latter per SWiTCH protocol (Helton, Blood 2014). Adjudication of differing reads was performed by a blinded third neuroradiologist. Participants undergoing MRI intentionally included 29 without any abnormal findings on the three other evaluations. Plasma samples were examined for key cytokines and markers of vascular activation by a 20-plex Luminex immunoassay. Immune markers included IL1b, IL1RA, IL2, IL4, IL10, IL18, IL22, IFNg, TNFa, TNFb, sFasL, CXCL1, CXCL8 (IL8), PDGFBB, VEGFA, serpin E1 (Plasminogen Activator Inhibitor (PAI)-1), sICAM1, VCAM1, TGFa, plus assay of hsCRP. Raw immune data were transformed using Box-Cox transformations and standardized; markers with persistently skewed (non-Gaussian) distributions after transformation were dichotomized for analysis. Continuous immune variables were analyzed by repeated measures ANOVA (unadjusted) and ANCOVA (adjusted for baseline covariates of age, sex, hemoglobin and 25-hydroxy-vitamin D3/D2. Dichotomized biomarkers were analyzed using generalized linear mixed-effects models. For these exploratory biomarker discovery-focused analyses, all significance tests were performed 2-tailed without adjustment for multiple comparisons, with a=0.1 for main effects. Results: Mean age was 6.48 ± 2.75 years, 50.6% male. Mean hemoglobin was 7.26±0.90 g/dl; 16.7% were malnourished using standard international measures established for age and sex. Infarcts on MRI were detected in 42 (52%), including 13 (25%) with no other test abnormalities. Of the 20 inflammatory markers tested, the levels of 13 markers were associated with one or more abnormality. One or more infarcts on MRI as the sole abnormality ("silent" stroke) were associated with levels IL1b, IL2 and TGFb (Table 1). Neurocognitive dysfunction alone was associated only with levels of IL22. Abnormal TCD (primarily "conditional" values) or prior stroke was associated with levels of VEGFA, IL4 and TNFb. In contrast, abnormal TCD or stroke by examination plus neurocognitive dysfunction was associated with nine cytokines levels: CXCL8, CXCL1, IFNg, TNFa, sFasL, IL18, IL22, sICAM1 and VEGFA. In all cases, cytokine levels were negatively associated with each of the abnormal neurological, neurocognitive or imaging results. Adjusting for age, sex, hemoglobin and malnutrition had only modest effects on statistically significant results. No associations were found between the test results and any of the other eight inflammatory markers, including hsCRP. Conclusions: In this exploratory biomarker analysis in a sample with highly prevalent abnormal test results, the presence of those abnormalities correlated with levels of a majority of the inflammatory markers assayed. The clinically well status at sampling and the finding of lower markers levels associated with existing abnormalities suggest that these children were not currently inflamed. As a cross-sectional analysis, cerebral vasculopathy underlying those abnormalities had previously occurred. Lower levels of inflammation may reflect a survivor status of children affected by a chronic illness with high local mortality. Our findings need to be confirmed in a prospective study children, and may be most interesting at acute events and for subjects receiving disease-modifying therapy. Disclosures No relevant conflicts of interest to declare.

Description: Introduction. Plasmodium falciparum malaria is a major cause of morbidity in African children with sickle cell anemia (SCA). Factors associated with severe disease and mortality in malaria, including tumor necrosis factor (TNF)-alpha and components of the angiopoietin (Angpt)-Tie-2 system, have also been implicated in the pathogenesis and clinical severity of SCA. However, there is no data on how these factors are altered in children with SCA during severe malaria. Methods. A total of 232 children who presented with severe malarial anemia (hemoglobin 〈 5 g/dL with Plasmodium parasitemia on peripheral blood smear) were enrolled in a prospective study of severe malaria at Mulago National Referral Hospital in Kampala, Uganda. No child had known SCA at the time of severe malarial anemia diagnosis. Samples from enrolled children were subsequently tested by genotyping for the presence of hemoglobin S (HbS) using a TaqMan assay at rs334. Clinical and laboratory parameters, plasma markers of inflammation and endothelial activation, and the estimated total, sequestered, and circulating parasite biomass were compared in children with HbSS compared to HbAA. Results. The study cohort included 208 children with HbAA (90.4%), 22 children with HbSS (9.6%), and 2 with HbAS. Children with HbSS were older than children with HbAA (Table 1), so all comparisons were adjusted for age. Children with HbSS versus HbAA did not differ significantly in duration of symptoms, clinical signs, disease severity, or degree of peripheral P. falciparum parasitemia. However, children with HbSS had significantly lower concentrations of PfHRP2, a marker of total parasite biomass, and lower levels of estimated sequestered parasite biomass (Table 1). Children with HbSS had pronounced leukocytosis, a feature of chronic inflammation in SCA, but had significantly lower concentrations of the inflammatory biomarkers C-reactive protein and alpha-1-acid glycoprotein and the pro-inflammatory cytokine TNF-alpha than children with HbAA (Table 1). In contrast, concentrations of angiopoietin-2 (Angpt-2), a marker of endothelial dysregulation that has been associated with mortality in severe malaria, were 3-fold greater in children with HbSS than HbAA (Table 1), and were associated with an increased risk of post-discharge recurrent malaria in the cohort, after adjustment for age, sex, and hemoglobin S genotype (odds ratio per log-10 increase in Angpt-2 [95% confidence interval], 2.11 [1.01, 4.38]). Conclusion. In this population, undiagnosed SCA is common in children with severe malarial anemia. During episodes of severe malarial anemia, children with SCA suppress parasite sequestration and inflammation but upregulate Angpt-2, which may increase risk of recurrence of malaria. Disclosures Conroy: ALC: Patents & Royalties: angiopoietin-1, angiopoietin-2. Ware:Addmedica: Other: Research Drug Donation; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Nova Laboratories: Membership on an entity's Board of Directors or advisory committees; CSL Behring: Membership on an entity's Board of Directors or advisory committees; Novartis: Other: DSMB; Bristol Myers Squibb: Other: Research Drug Donation.