Painful Vaso-occlusive Crises Research Articles

Contribution of Vascular Cell Adhesion Molecule to Hemodynamics in Sickle Cell Disease

Vaso-occlusive events (VOEs) and pain crises are common clinical features of sickle cell disease (SCD) that result from sickle-shaped erythrocytes and leukocytes blocking blood flow, particularly in small vessels (Kato 2018). Activated endothelium also plays a role in the pathogenesis of VOEs in SCD. For example, VCAM-1 is expressed on blood vessels after activation by chemical and/or mechanical stimulation, which results in cytokine release. Studies have shown that patients with SCD have higher steady-state serum levels of soluble VCAM-1 compared to controls and that these levels are elevated during VOEs (White 2020). Furthermore, overexpression of these adhesion molecules on the endothelium results in prolonged adherence of white blood cells (WBCs), which has been shown to contribute to the development of VOEs and possibly cerebral vasculopathy. These findings raise the need to explore further the role of aberrant WBC- and/or RBC-endothelial interaction, mediated via VCAM-1, in the pathophysiology of cerebral microvascular hemodynamics and vasculopathy leading to cerebral microinfarcts in SCD. Therefore, we hypothesized that sickle cell mice will show greater cerebral cortical expression of VCAM-1 compared with age-matched controls and that this deposition will be associated with significant evidence of abnormal cerebral microvascular hemodynamic abnormalities.To examine the relationship between abnormalities in cerebral microvascular hemodynamics and VCAM-1 deposition in the cerebral microvasculature, we utilized a humanized sickle cell (with HbSS) and corresponding control (with HbAA). After cranial-window procedures, cortical capillaries, precapillary arterioles, and post-capillary venules were imaged using two-photon microscopy at two time points. In addition, this experiment included pre-and post-transfusion groups as we intend to study the impact of blood transfusion on hemodynamics. Using custom-written but well-validated MATLAB scripts, we analyzed line scans to identify the number and duration of rolling or adherent WBCs and RBCs, the RBC velocity in cerebral microvasculature, and the frequency and magnitude (mL/sec) of cerebral microvascular blood flow reversals. Rolling WBCs were defined as lasting two seconds or more, and adherent RBCs were defined as lasting 0.5 seconds or more. To quantify the expression of VCAM-1, we used immunohistochemistry to stain 50-micron sections of brain tissue for VCAM-1, Lectin to localize the vasculature, and Neun to localize neuronal nuclei. Images were analyzed using Phenochart and ImageJ software to examine the deposition of VCAM-1 throughout the brain tissue.As shown in Figure 1, at the first time point (baseline), we observed a significantly higher maximum RBC velocity (p<0.001) in the sickle cell mice compared to controls (figure 1a). We also found that there was significantly higher expression of VCAM-1 (p<0.001) (figure 1b) as well as significantly more leukocyte rolling (p<0.001) (figure 1c) in the sickle cell mice compared to controls. Additionally, we noted that the sickle cell mice have a significantly higher frequency of blood flow reversals (p<0.01) (figure 1d) as well as higher magnitude of microvascular blood flow reversals (p<0.001) (figure 1e) compared to controls. Interestingly, the sickle cell mice have a slightly lower average or mean capillary blood flow velocity compared to control (figure 1f), but this was not statistically significant (p=0.079). Since the mean capillary velocity is obtained as a smoothened difference between the forward flow and reversals, this decrease was surprising given the significant differences in frequency and magnitude of microvascular blood flow reversal in the sickle cell mice compared to controls (figs 1d and 1e).In conclusion, we see that the high velocity of blood flow might be a mechanical force, among other factors contributing to cerebral microvascular VCAM-1 expression in sickle cell mice. This might be responsible for the increased leukocyte-endothelial interactions and adhesion, ultimately leading to higher frequency and magnitude of cerebral microvascular blood flow reversal. Taken together, this may contribute to the observed slightly lower mean or effective microvascular forward blood flow. These pathophysiological changes might contribute to the reported higher rate of cerebral microinfarct and silent infarct in sickle cell disease. [Display omitted] DisclosuresArcher: Global Blood Therapeutics: Consultancy, Research Funding; Forma Therapeutics: Research Funding. Hyacinth: Novartis: Consultancy; Acuta Capital: Consultancy.

Sickle Cell Disease with Dermatomyositis - a Rare and Complex Comorbidity

Introduction and Case Presentation - A 4 yrs old female with sickle cell disease (SCD) and intermittent asthma presented with polyarthralgia predominantly involving bilateral hip and knee joints and became non-ambulatory over a course of 2 months. She developed chronic facial swelling, and a pruritic erythematous rash involving face, extensor surface of the hand and the right knee with significant weight loss. Physical examination was significant for heliotropic rash on the eyelids, Also present were Gottron's papules and macules on both hands and right knee along with right leg tenderness, muscle weakness, wasting and decreased range of motion at bilateral hip joints along with inguinal lymphadenopathy. Investigations were significant for severe anemia with hyperactive bone marrow, metabolic acidosis, normonatremic dehydration with absence of any evidence of an ongoing infectious process. Her Anti Nucleic acid antibody (ANA) panel, Aldolase, Rheumatoid factor and Angiotensin-1-Converting Enzyme levels were unremarkable. Creatine kinase levels were near normal. Her Neopterin and LDH levels were significantly elevated. Imaging with MRI scan with multiplanar multisequence of bilateral lower extremities and Pelvis showed significant diffuse myositis and soft tissue swelling. The diagnostic criterion for Juvenile Dermatomyositis include: (1) the typical pathognomonic rashes, (2) elevated muscle enzymes, (3) symmetric proximal muscle weakness and neck flexors, (4) muscle biopsy characteristic of juvenile dermatomyositis, and (5) EMG findings characteristic of juvenile dermatomyositis. Having ruled out an infectious process a muscle biopsy was performed, which also was subjected to Electron Microscopic (EM) analysis as the patient did not have extremely high muscle enzymes, necessitating a muscle biopsy . Hematoxylin & eosin stained sections were compatible with inflammatory myopathy. Ultrastructural examination demonstrated myofiber size variability, frequent rounded atrophic fibers with myofibrillar disarray, internalized nuclei, and increased interstitial collagen deposition. Endomysial capillaries were decreased in number, but showed reactive endothelial changes. Some showed prominent endothelial tubuloreticular inclusions (Figure),characteristic of JD. In summary, absence of serologic and tissue evidence of any other inflammatory myopathy, offending pharmacotherapy, infectious disease and presence of imaging and tissue evidence (vascular injury and tubuloreticular inclusions) comprised the work up of a child with SCD with Dermatomyositis.Management- Prompt treatment with prednisone (2 mg/kg/day), methotrexate (10 mg) weekly and monthly IVIG (2gm/kg/dose) infusions was started, once diagnosis was confirmed showing a dramatic clinical response. Prednisone was slowly tapered after 1 month of treatment. She showed gradual improvement in her symptoms, the strength in proximal muscle in both upper and lower limbs improved eventually to a point where the contractures in the elbows and knees improved and she started ambulating without support. Patient's Hydroxyurea was restarted at a low dose of 15mg/kg/day and increased to 20 mg/kg/day slowly, as her systemic symptoms of Dermatomyositis subsided.Discussion- Working up a child with SCD who presents with sudden onset non weight bearing can be complicated. Vaso-occlusive bony pain crisis , Osteomyelitis , septic arthritis are common clinical scenarios, but an underlying rheumatologic illness in pediatric SCD patient mimicking a sickle cell pain crisis presents a unique diagnostic challenge one which, albeit has been reported in literature, but rarely includes electron microscopic ultrastructural examination as part of evidence and work up. This patient's relatively mild muscle enzyme elevation and Hydroxyurea therapy made the initial diagnosis more challenging. We present the first pediatric SCD patient with clinically and pathologically proven case of Dermatomyositis with EM evidence, highlighting a unique clinical scenario ,diagnostic challenges and management strategies. [Display omitted] DisclosuresJain: Octapharma: Consultancy; CSL Behring: Consultancy, Speakers Bureau; GBT: Consultancy; Blue Bird Bio: Consultancy; Takeda: Consultancy, Speakers Bureau.

Quality of Life in Children, Adolescents and Young Adults with Sickle Cell Disease and Their Caregivers during Standard of Care and after Bone Marrow Transplantation: A Single Center Report

▪BackgroundIn spite of the improvements in clinical care of children with Sickle Cell Disease (SCD), painful vaso-occlusive (VOC) crises, recurrent admissions and long hospital stays contribute to the disruption of the social and school life of children and adolescents with SCD causing a poor QoL.Limited information is available regarding QoL of children, adolescents and young adults with SCD and their caregivers during standard of care and after bone marrow transplantation in Italy even though Italian patients participating in international meetings or global surveys highlighted the importance to improve QoL (Strunk C. BMC Proc. 2020, Osunkwo I. Am J Hematol. 2021). Moreover, no mention is given to QoL in the current AIEOP Recommendations for the Management of Children with SCD in Italy. The availability of new treatment options for SCD highlights the need to improve QoL evaluation before and after treatments.Our Center decided, therefore, to include QoL evaluation as part of comprehensive care for patients with SCD. This study has the following aims: to describe the QoL of children, adolescents and young adults with SCD undergoing standard care or after disease curative treatments (bone marrow transplantation) and the QoL of their caregivers; to evaluate the correlation of QoL with clinical-haematological and therapeutic variables.MethodsHealth Related QoL was examined with the Patient-Reported Outcomes Measurement Information System (PROMIS) questionnaires in Italian: Parent Proxy Profile-49 v2.0, Pediatric Profile-49 v2.0 and 57 Profile v2.1, exploring 8 domains: Pain, Fatigue, Anxiety, Depression, Physical Abilities, Peer Relations, Sleep Disorders, Pain Interference. An English version was available for English speaking parents.Patients and caregivers accessing the SCD Clinic starting May 2021 were given the paper version of the questionnaires; due to the COVID pandemic and the limited access to the SCD Outpatient Clinic, a link to a Google online version of the questionnaires was provided to all teenagers and young adults, through their mobile devices. .PROMIS Scores were standardized through the Health Measures Scoring Service (healthmeasures.net). For the descriptive analysis, the T-score was obtained, for each patient and for each PROMIS domain (symptom or function), classifying impairment in each domain as normal, mild, moderate, or severe. The Student T Test for comparisons of the means among samples and the Wilcoxon Test for the sum of ranks were used in the statistical analysis of normal and non-normal continuous variables. For the correlation analysis between continuous variables of which at least one is not normal, the Spearman Correlation Test was used. The values with p<0.05 were considered statistically significant.ResultsAll patients and parents approached accepted to perform the questionnaire. The study involved 18 caregivers and 50 patients (25% F, mean age 16.4 years, 74% HbSS, 76% from Africa): 41 undergoing standard care (7 no therapy, 34 Hydroxyurea or chronic transfusion) and 9 who received HSCT. 37 patients (74%) and 8 parents (44%) completed the online Google version of the questionnaire. The standard of care patients displayed mild to severe symptoms in various domains (Figure 1A); in the transplant population there was impairment in QoL, with less severe impairment in most of the domains, especially in the pain domains, than what was in the standard of care group. (Figure 1B).Anxiety levels and depressive symptoms were greatest between the ages of 14-26, compared to younger ages (p 0.018).Parents do not have the same perception of the disease as their children: they appeared to overestimate the domain of pain and fatigue and underestimate anxiety and depression (p <0.001). Sleep quality was impaired in both affected and HSCT patients.The number of hospital admissions in the previous year correlated with worse QoL (p 0.04), while the number of painful VOC showed a tendency towards significance (p 0.07); there was no difference with the other domains. Updated results will be presented.ConclusionsOur data show the feasibility of evaluating QoL during routine visits and also remotely. Impairment of QoL is already present in a subgroup of young patients. Even after HSCT, QoL is not optimal but personal, social, and economic reasons need to be taken into account to adequately interpret the results. Longitudinal assessment to look at QoL will be important. [Display omitted] DisclosuresBiffi: BlueBirdBio: Consultancy, Other: Advisory Board. Colombatti: Global Blood Therapeutics: Consultancy; BlueBirdBio: Consultancy; NovoNordisk: Consultancy; Novartis: Consultancy; Forma Therapeutics: Consultancy; Addmedica: Consultancy; Global Blood Therapeutics: Research Funding; BlueBirdBio: Research Funding.

Hyperhemolysis Syndrome in SCD Patient: Reminder of a Rare but Life-Threatening Complication of Blood Transfusion

Background: Sickle cell disease (SCD) patients are at risk of developing multiple complications from transfusions, including alloimmunization to red blood cell (RBC) antigens, delayed hemolytic transfusion reactions, and hyperhemolysis syndrome (HS). HS is a serious complication of transfusion characterized by the destruction of both transfused and autologous RBCs with resulting severe anemia and post transfusion hemoglobin lower than pretransfusion levels. We report the case of a middle age female patient with known SCD who developed severe HS following a blood transfusion. We aim to remind physicians of the importance of conservative blood transfusions in SCD patients in order to avoid serious transfusion-related complications.Case report: A 57-year-old African American patient, with known history of SCD who was doing well with a baseline hemoglobin (Hgb) of 6-7 g/dl. Transfusion history included 4 units of Packed Red Blood Cell (PRBC) during the 5 years prior to this presentation, all of which for mild, non-resolving vaso-occlusive pain crisis. Her most recent transfusion was 7 days prior to her presentation, she received 1 unit of PRBC for a Hgb level of 6.3 g/dl, associated with mild musculoskeletal pain and fatigue. She presented to the Emergency Department 4 days later with worsening fatigue, decreased oral intake and dark urine. On presentation, she was normotensive, afebrile and mildly tachycardic. She had increasing oxygen requirements to maintain O2 saturation above 94%. Her blood work showed a Hgb of 2.8 g/dl (12-15 g/dL), hematocrit 8.3 % (36-46 %), RBC count 0.87 M/uL (4.15-5.55 M/uL), Mean Corpuscular Volume 95.5 fl (80-100 fl), elevated White Cell Count at 28.4 K/uL (3.8-10.6 K/uL), and platelet count 125 K/uL (150-450 K/uL). Hemolysis labs showed low haptoglobin of < 30 mg/dl (30-200 mg/dl), elevated Lactate Dehydrogenase at 3420 IU/L (< 250 IU/L), total bilirubin 2.7 mg/dl (< 1.2 mg/dl), direct bilirubin 0.6 mg/dl (0-0.3 mg/dl), and reticulocyte count 3.5% (0.5-1.5 %; reticulocytopenia relative to degree of anemia). A disseminated intravascular coagulation (DIC) panel showed fibrinogen of 263 mg/dL (200-450 mg/dL), D-dimer greater than 20 ug/mL (< 0.50 ug/ml), prothrombin time of 19.8 seconds (s) (11.5-14.5 s), and partial thromboplastin time of 32 s (22-36 s). High sensitivity troponin was elevated at 650 ng/L (< 19 ng/L). Antibody screen and direct antiglobulin test (DAT) were negative. Peripheral blood smear showed severe anemia with marked anisopoikilocytosis including numerous blister cells, occasional sickle cells and numerous nucleated red blood cells. The recent history of blood transfusion and the current laboratory workup were consistent with HS.Patient was admitted to the intensive care unit (ICU) for management; she initially received 1g intravenous iron dextran and intravenous immunoglobulin (IVIG) 0.4 g/kg for 5 days. She was also started on erythropoietin, folic acid, and vitamin B12. Her reticulocyte count improved to 19%. Given no improvement in Hgb levels, systemic steroids were started after ruling out infectious etiologies. She initially received methylprednisolone 125mg daily for 2 days, followed by oral prednisone 60mg daily for 7 days. Patient had increased oxygen requirements during admission, had an elevated lactate to 4 mmol/L, and had a drop in Hgb to 2.1 g/dL. She was still managed conservatively with oxygen supplementation and intravenous crystalloid fluids. The decision was to avoid transfusions unless they were life-saving. Patient remained in the ICU unit for 5 days, then was transferred to the hematology floor where she remained hospitalized for 7 days. Oxygen requirements and patient's symptoms steadily improved, hemolysis labs trended down, and reticulocyte count improved. Hgb levels improved gradually to highest of 5.7 g/dl prior to discharge. Patient was then discharged to follow up with her hematologist in the outpatient setting.Conclusion: This case aims to highlight the importance of early recognition of HS to avoid wrong management with RBC transfusion. Our patient had severe anemia and was managed with transfusion-free approach with good outcome. This case is also meant to remind physicians of the importance of conservative blood transfusions in SCD patients in order to avoid serious and life-threatening transfusion-related complications. DisclosuresNo relevant conflicts of interest to declare.

Effects of Quercetin on Neutrophil Extracellular Trap Formation in Sickle Cell Disease

Introduction: Sickle Cell Disease (SCD) is characterized by chronic inflammation with innate immune cell activation, especially observed in neutrophils. Emerging evidence implicates the imbalance between neutrophil extracellular trap (NET) formation and degradation as having a central role in the pathophysiology of thromboinflammation and venous thrombosis. Although NETosis and NET formation influences venous thromboembolism (VTE) pathophysiology, little is known about baseline and agonist-induced NETosis in SCD. We hypothesized that systemic neutrophil activation would lead to higher baseline and agonist induced NETosis in SCD and would influence phenotypic variability. To test this hypothesis, we assessed baseline and agonist induced NETosis in patients with SCD and ethnic matched controls. We also investigated the anti-inflammatory effects of flavonoid Quercetin on neutrophil activation.Methods: Neutrophils negatively selected from citrate anticoagulated blood using an immunomagnetic bead based kit (MACSxpress® Miltenyi Biotec) were either fixed immediately to assess baseline NETosis or stimulated with fMLP (1 µM) for 1 hour to assess agonist-induced NETosis. To study flavonoid anti-inflammatory effects, neutrophils were pretreated with Quercetin (100 µM) for 30 min prior to fixation and fMLP stimulation. NETosis was assessed by flow cytometry. Extracellular DNA extrusion on neutrophils was detected by gating the neutrophil population staining with Sytox green. Sytox green positive neutrophils that were positive for both myeloperoxidase (MPO) and tri-Citrullinated Histones (H3Cit) were defined as undergoing NETosis. In some experiments, NET formation was independently confirmed by image flow cytometry (AMNIS).Results: Subjects included SCD patients (genotype SS n=11) and ethnic matched controls (genotype AA, n=11) with a median age of 49 years (p=0.58) and a predominance of males (70%). All SCD patients were at least 60 days remote from an acute painful vaso-occlusive crisis or blood transfusion and were receiving hydroxyurea. The white cell and absolute neutrophil counts were higher in SCD patients (mean ± SD 8.77 ± 1.52 and 5.07 ± 1.78 x 10 9/L) when compared with controls (mean ± SD 5.33 ± 1.05 and 2.8 ± 0.95 x 10 9/L). Subsequent data are presented as median percentages with interquartile ranges (IQR).A subgroup of the study population demonstrated spontaneous NETosis (27%; SS = 4; AA = 4) and were therefore excluded from our analysis. Contrary to expectations, SCD patients exhibited a lower percentage of NETosis at baseline compared to controls (20 % (11, 36) vs. 33 % (15, 58); p=0.22). Similarly, neutrophils from SCD patients exhibited lower agonist-induced NETosis compared to controls (42% (19, 47) vs. 51% (37, 70); p=0.15) (Fig 1 A and B)Pretreatment of neutrophils from SCD patients with Quercetin appeared to inhibit basal levels of NETosis (6%, (2, 26) vs. 20% (11, 36) p=0.08) although this effect was not appreciable in controls (33% (11, 58) vs. 33% (15, 58) p=0.41) (Fig 1 C and D). Neutrophils from SCD patients that were pretreated with Quercetin and then stimulated with fMLP demonstrated significantly reduced NETosis compared to untreated neutrophils (17.1% (10, 38) vs 41.7% (19, 47) p=0.007) although this effect was not significant in controls (35% (17, 72) vs 50.7% (37, 70) p=0.11) Fig 1 E and F.Our ongoing experiments will demonstrate the effects of more specific inhibitors of neutrophil activation (e.g. R406) in human and mouse models of SCD.Conclusion: These preliminary data suggest lowered NETosis in SCD patients despite neutrophil activation in the systemic inflammatory environment that are partially explained by hydroxyurea treatment. The results also support further evaluation of anti-inflammatory therapies to reduce neutrophil activation in SCD and ameliorate thrombo-inflammatory disease pathology. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Treatment with IMR-687, a Highly Selective PDE9 Inhibitor, Increases HbF and Reduces VOCs in Adults with Sickle Cell Disease in a Long-Term, Phase 2a, Open-Label Extension Study

IntroductionIMR-687 (tovinontrine) is a highly selective phosphodiesterase 9 (PDE9) inhibitor being developed as an orally administered therapy for patients with sickle cell disease (SCD) and beta-thalassemia. IMR-687 increases intracellular cGMP levels and has been shown in preclinical studies to increase fetal hemoglobin (HbF) expression and reduce hemolysis and sickling of RBCs, which can lead to painful vaso-occlusive crisis (VOC). In a Phase 2a, randomized, double-blind, placebo-controlled study of adult patients with SCD (N=93) (NCT03401112), IMR-687 was generally well-tolerated as a monotherapy and in combination with hydroxyurea (HU) (European Hematology Association Annual Congress 2021, Abstract S263). IMR-687 treatment with 50-200 mg once daily (N=63) for up to 6 months, as compared with placebo (N=30), resulted in a 40% lower mean annualized rate of VOCs, 38% lower mean annualized rate of VOC-related hospitalizations, and increased median time to first VOC (169 days vs 87 days, respectively, p=0.029). Improvements were also observed in patient-reported outcomes regarding pain episode severity (ASCQ-Me) and percentage of RBCs containing HbF (F-cells).MethodsA Phase 2a open-label extension (OLE) study (NCT04053803) is ongoing to assess the long-term safety and benefit of IMR-687 administered to adult subjects with SCD for up to 4 years. Secondary long-term pharmacokinetic (PK) and pharmacodynamic (PD) parameters are also being examined. The use of background HU is permitted. The Safety Review Committee meets regularly and ad hoc as necessary.Subjects were eligible upon completion of the Phase 2a parent study (16 or 24 weeks depending on the assigned treatment arm and without early withdrawal) and if they met all inclusion and exclusion criteria. For subjects without treatment interruption, baseline values from the parent study were used, as appropriate. The dose of IMR-687 initially administered in the OLE study was 200 mg as a once daily oral dose. The dose of IMR-687 is planned to be increased to 300 or 400 mg daily, depending on the subject's weight.Subjects are evaluated every 4-6 months for safety, PK and PD parameters (including HbF, F-cells, hemoglobin [Hb] and other biomarkers) as well as incidence of VOCs.ResultsAs of a data cut-off of 12-May-2021, 24 subjects were enrolled in the OLE study, 17 of which had a treatment interruption between the parent and OLE studies, and 7 subjects had direct roll-over to the OLE study. Seventeen of the OLE subjects were treated with IMR-687 monotherapy and 7 subjects were treated with combination IMR-687 + HU.IMR-687 continued to be well-tolerated in the OLE study (N=24). There were no treatment-related serious adverse events (SAEs), and the most common (≥10%) AEs were headache (21% of subjects), back pain (17%) and nausea (13%). There were 3 (13%) subjects with VOC-related hospitalizations.Annualized VOC rate was analyzed for 18 subjects treated for at least 200 days in the OLE study. Subjects who were previously in the active treatment arm in the parent study (N=13) maintained a low mean annualized VOC rate while continuing IMR-687 in the OLE study (1.35/yr [parent] and 1.85/yr [OLE]); subjects previously treated with placebo (N=5) had a 39% reduction in VOC rate when switched to IMR-687 (4.71/yr [parent] vs 2.89/yr [OLE]) (Figure 1). Of 15 subjects with evaluable PD data at 8 months, 7 (47%) subjects had a ≥ 6% absolute increase in F-cells, and 4 of 11 (36%) subjects had a ≥ 3% absolute increase in HbF (Figure 2). Updated data for subjects completing 12 months in the OLE study will be presented.ConclusionsConsistent with the parent study, preliminary results from the ongoing Phase 2a OLE study demonstrate that daily dosing of 200 mg IMR-687 was well-tolerated with longer-term treatment as a monotherapy or in combination with HU. IMR-687 treatment reduced the annualized rate of VOCs and increased HbF (%) and F-cells (%). Based on these encouraging data, a Phase 2b study (NCT04474314) is ongoing to further explore IMR-687 at doses up to 400 mg daily as a disease-modifying therapy for SCD. [Display omitted] DisclosuresBarysauskas: Imara Inc.: Current Employment, Current equity holder in publicly-traded company. Yen: Imara Inc.: Current Employment, Current equity holder in publicly-traded company. Tang: Imara Inc.: Current Employment, Current equity holder in publicly-traded company. Ballal: Imara Inc.: Current Employment, Current equity holder in publicly-traded company. Attie: Imara Inc.: Current Employment, Current equity holder in publicly-traded company.

PEDIATRIC PLASTIC BRONCHITIS IN THE SETTING OF SICKLE CELL DISEASE TREATED WITH INHALED TISSUE PLASMINOGEN ACTIVATOR

TOPIC: Pediatrics TYPE: Medical Student/Resident Case Reports INTRODUCTION: Plastic bronchitis is seen in congenital heart disease, cystic fibrosis, asthma, and less commonly sickle cell disease. It has high morbidity and can be difficult to treat. CASE PRESENTATION: We present an 11-year-old African American boy with past medical history of hemoglobin SS disease, acute chest syndrome, pain crises, and obstructive sleep apnea status post tonsillectomy and adenoidectomy who presented with vaso-occlusive pain crisis consisting of chest pain. CXR was clear without consolidation. He was placed on a pain regimen of morphine and ketorolac. Acute chest prophylaxis was initiated with albuterol plus PEP every 4 hours, incentive spirometry, and oxygen saturation maintenance ≥95%. Over the next two days he developed fevers, increased oxygen requirement, and crackles. CXR progressed to consolidation of left lung. He was transferred to the Pediatric ICU for BiPAP and given 2 units pRBCs, 7 days ceftriaxone, and 5 days azithromycin for acute chest treatment. Despite intense respiratory treatments while on BiPAP with albuterol every 4 hours, dornase alfa every 12 hours, CPT every 4 hours, and home mometasone/formoterol every 12 hours, his CXR demonstrated persistent opacification of left lung. Pulmonology was consulted for bronchoscopy which demonstrated large white mucoid material in the left mainstem consistent with plastic bronchitis. The cast was unable to be removed on initial bronchoscopy. He was initiated on inhaled tissue plasminogen activator 4 times daily in addition to above respiratory treatments. Bronchoscopy was repeated two days later and cast was successfully removed. BiPAP and oxygen were weaned as tolerated and he was discharged home on mometasone/formoterol twice daily. DISCUSSION: Plastic bronchitis describes endobronchial plugs of rubber-like casts in the bronchial trees. It is more commonly seen in patients with congenital heart disease after single-ventricle palliation. There have been various treatments trialed including bronchoscopy, chest physiotherapy, inhaled steroids, hypertonic saline, dornase alfa, nebulized heparin, lymphatic embolization, and inhaled tissue plasminogen activator. Studies performed on casts have demonstrated a substantial fibrin component supporting inhaled tissue plasminogen activator as a potentially effective treatment option. CONCLUSIONS: We present a case of pediatric plastic bronchitis in the setting of a patient with sickle cell disease. This case emphasizes that plastic bronchitis must be considered in patients diagnosed with acute chest. Additionally, inhaled tissue plasminogen activator may be considered as a treatment modality in pediatric patients with plastic bronchitis, particularly in patients who are unable to undergo bronchoscopy or when it has been unsuccessful for removal. REFERENCE #1: Feray S, Mora P, Decavele M, Pham T, Hafiani EM, Fartoukh M. Plastic bronchitis: An unusual complication of acute chest syndrome in adult. Respir Med Case Rep. 2017 Apr 7;21:93–5. REFERENCE #2: Li Y, Williams RJ, Dombrowski ND, Watters K, Daly KP, Irace AL, et al. Current evaluation and management of plastic bronchitis in the pediatric population. Int J Pediatr Otorhinolaryngol. 2020 Mar 1;130:109799. REFERENCE #3: Heath L, Ling S, Racz J, Mane G, Schmidt L, Myers JL, et al. Prospective, Longitudinal Study of Plastic Bronchitis Cast Pathology and Responsiveness to Tissue Plasminogen Activator (tPA). Pediatr Cardiol. 2011 Dec;32(8):1182–9. DISCLOSURES: No relevant relationships by Megan Lilley, source=Web Response No relevant relationships by Carla Roberts, source=Web Response No relevant relationships by Heather Staples, source=Web Response

Extracellular Vesicles in Sickle Cell Disease: Plasma Concentration, Blood Cell Types Origin Distribution and Biological Properties.

Prototype of monogenic disorder, sickle cell disease (SCD) is caused by a unique single mutation in the β-globin gene, leading to the production of the abnormal hemoglobin S (HbS). HbS polymerization in deoxygenated condition induces the sickling of red blood cells (RBCs), which become less deformable and more fragile, and thus prone to lysis. In addition to anemia, SCD patients may exhibit a plethora of clinical manifestations ranging from acute complications such as the frequent and debilitating painful vaso-occlusive crisis to chronic end organ damages. Several interrelated pathophysiological processes have been described, including impaired blood rheology, increased blood cell adhesion, coagulation, inflammation and enhanced oxidative stress among others. During the last two decades, it has been shown that extracellular vesicles (EVs), defined as cell-derived anucleated particles delimited by a lipid bilayer, and comprising small EVs (sEVs) and medium/large EVs (m/lEVs); are not only biomarkers but also subcellular actors in SCD pathophysiology. Plasma concentration of m/lEVs, originated mainly from RBCs and platelets (PLTs) but also from the other blood cell types, is higher in SCD patients than in healthy controls. The concentration and the density of externalized phosphatidylserine of those released from RBCs may vary according to clinical status (crisis vs. steady state) and treatment (hydroxyurea). Besides their procoagulant properties initially described, RBC-m/lEVs may promote inflammation through their effects on monocytes/macrophages and endothelial cells. Although less intensely studied, sEVs plasma concentration is increased in SCD and these EVs may cause endothelial damages. In addition, sEVs released from activated PLTs trigger PLT-neutrophil aggregation involved in lung vaso-occlusion in sickle mice. Altogether, these data clearly indicate that EVs are both biomarkers and bio-effectors in SCD, which deserve further studies.

A Randomized Clinical Hypnosis Pilot Study: Improvements in Self-Reported Pain Impact in Adults with Sickle Cell Disease.

Sickle cell disease (SCD) is characterized by recurrent painful vasoocclusive crises. Current evidence focuses on the frequency of acute pain crises resulting in emergency department use and nonplanned inpatient hospital admissions; yet few studies focus on pain sequelae outside the healthcare system or how individuals self-manage their chronic SCD-related pain. This study investigated the feasibility of a biobehavioral intervention as an adjunct nonpharmacological therapy to assist in the self-management of chronic pain. A randomized, controlled clinical trial of hypnosis was conducted in outpatients with SCD (n = 31). Patient-reported outcomes (PROs) administered at baseline, five, and twelve weeks from both groups included pain frequency, intensity, and quality (Pain Impact Scale (PIQ) and Numerical Rating Scales); anxiety (State-Trait Anxiety Inventory), coping strategies (Coping Strategies Scale), sleep (Pittsburgh Sleep Quality Index (PSQI)), and depression (Beck Depression Inventory (BDI)). The same PROs were collected at weeks seventeen and twenty-four from the control group after the crossover. No significant group by time interaction effects were found in any of the PROs based on the repeated-measures mixed models. The PIQ and PSQI scores decreased over time in both groups. Post hoc pairwise comparisons with the Bonferroni adjustment indicated that the mean PIQ score at baseline decreased significantly by week 12 (p = 0.01) in the hypnosis group. There were no significant changes across time before and after the crossover in any of the PROs in the control group. As suggested by these findings, pain impact and sleep in individuals with SCD may be improved through guided mind-body and self-care approaches such as hypnosis.

Indirect Economic Burden of Sickle Cell Disease

ObjectiveThis study aimed to quantify the indirect costs of sickle cell disease in the United States. MethodsAdult patients from a sickle cell disease clinic at an urban academic healthcare system completed an adapted Institute for Medical Technology Assessment Productivity Cost Questionnaire related to the impact of their disorder on absenteeism, presenteeism, ability to contribute through unpaid work outside of employment, and other aspects of life. Additional data were collected from patient records about each participant’s genotype, total hemoglobin level, and pain level. ResultsOf the 192 individuals, 187 who completed the survey reported experiencing vaso-occlusive crisis pain events during the last year that negatively affected their productivity at work and in daily roles. Three-fourths of respondents reported impairment in their ability to complete everyday tasks, such as caring for children, running errands, doing housework, shopping for groceries, and volunteer (unpaid) work. Only 30% of respondents reported being employed or self-employed. Of those employed, estimated costs of absenteeism and presenteeism attributable to pain events averaged $15 103 per person annually. Estimated total annual losses in unpaid work productivity averaged $3 145 862 for the study respondents and another $2 870 652 for their caregivers. ConclusionsSickle cell disease affected the work productivity, nonwork productivity, and the daily lives of adults seen with the disorder in an academic medical center.

Identifying elevated risk for future pain crises in sickle-cell disease using photoplethysmogram patterns measured during sleep: A machine learning approach.

Transient increases in peripheral vasoconstriction frequently occur in obstructive sleep apnea and periodic leg movement disorder, both of which are common in sickle cell disease (SCD). These events reduce microvascular blood flow and increase the likelihood of triggering painful vaso-occlusive crises (VOC) that are the hallmark of SCD. We recently reported a significant association between the magnitude of vasoconstriction, inferred from the finger photoplethysmogram (PPG) during sleep, and the frequency of future VOC in 212 children with SCD. In this study, we present an improved predictive model of VOC frequency by employing a two-level stacking machine learning (ML) model that incorporates detailed features extracted from the PPG signals in the same database. The first level contains seven different base ML algorithms predicting each subject's pain category based on the input PPG characteristics and other clinical information, while the second level is a meta model which uses the inputs to the first-level model along with the outputs of the base models to produce the final prediction. Model performance in predicting future VOC was significantly higher than in predicting VOC prior to each sleep study (F1-score of 0.43 vs. 0.35, p-value <0.0001), consistent with our hypothesis of a causal relationship between vasoconstriction and future pain incidence, rather than past pain leading to greater propensity for vasoconstriction. The model also performed much better than our previous conventional statistical model (F1 = 0.33), as well as all other algorithms that used only the base-models for predicting VOC without the second tier meta model. The modest F1 score of the present predictive model was due in part to the relatively small database with substantial imbalance (176:36) between low-pain and high-pain subjects, as well as other factors not captured by the sleep data alone. This report represents the first attempt ever to use non-invasive finger PPG measurements during sleep and a ML-based approach to predict increased propensity for VOC crises in SCD. The promising results suggest the future possibility of embedding an improved version of this model in a low-cost wearable system to assist clinicians in managing long-term therapy for SCD patients.

Effect of Immunozin™ therapy on hematological and biochemical parameters of Sickle Cell Disease patients who presented with fatigue, acute bone pain, and acute chest syndrome in Nigeria

Introduction: Painful vaso-occlusive crises are frequent complications of sickle cell disease (SCD), which affects about 3% of the Nigerian population with a high mortality in children. Relevance of hematological and biochemical parameters to clinical symptoms of SCD are not often documented. Aims and Objective: This study aimed at determining the prevalence of fatigue, acute bone pains (ABP) and acute chest syndrome (ACS) and to assess the associating hematological and biochemical parameters before and after administration of ImmunozinTM. Materials and Method: This study was a double-blind, two-arm, randomized control pilot study with 30 SCD patients presenting with fatigue, acute bone pains (ABP) and ACS, each of whom was given the study agent. At the first visit, after thorough assessment of each patient had been undertaken and hematological parameters were measured, the study drug was administered to each patient accordingly. This process was repeated monthly for five more monthly visits when the study concluded. At each visit, venous blood sample was collected for hematological parameters, electrolytes, urea and creatinine analyses within 2 hours of collection. Results: Means (±sd) of age and BMI (Kg/m2 ) of the study subjects were 13.0 (5.6) and 16.1 (1.9) respectively with 26 (86.7%) of the patients underweight. At enrollment into the study, 25 (83.3%), 18 (60.0%) and 10 (33.3%) of the patients presented with ABP, fatigue and ACS respectively but at the end of study, post administration of study drug, 11 (36.7%), 8 (26.7%) and 7 (23.3%) presented with ABP, fatigue and ACS. At enrollment, females were less likely to present with moderate fatigue (χ²=2.03, P-value=0.15, OR=0.23, 95% CI: 0.05, 1.18), moderate ABP (χ²=0.09, P-value=0.77, OR=0.80, 95% CI: 0.18, 3.46) and moderate ACS (χ²=0.01, P-value=0.90, OR=0.40, 95% CI: 0.03, 4.96) compared to males. Only one patient each presented with fatigue and ABP at the end of study. The values of many hematological parameters, electrolytes, urea and creatinine significantly varied at the end of the study compared with enrollment values. A significant positive correlation (Pearson’s r=0.40, P-value=0.031) was observed between serum urea and ACS at enrollment, and astonishingly between fatigue and ABP (r=0.52, P-value=0.003) but no notable correlation between any of the clinical symptoms and other variables at the end of study. Conclusion: The significant reduction in proportion of patients with fatigue, ABP and ACS at the end of study, after administration of test drug may suggest the therapeutic consequence of the test drug among SCD patients. There is urgent need to conduct multi-center and multi-disciplinary studies to corroborate these findings.

Enhanced Renal Clearance of Heme Triggers Acute Kidney Injury in Sickle Cell Disease

Circulating heme is scavenged primarily by hemopexin (Hx) for degradation in the liver, while alpha-1-microglobulin (A1M) binds heme for urinary clearance. Individuals with sickle cell disease (SCD) have acquired Hx deficiency. Hitherto, it has not been determined whether the low Hx in SCD results in a disproportionately large amount of heme exposure in the kidneys sufficient to promote acute tissue injury. Acute kidney injury (AKI) is a common downstream complication of acute chest syndrome (ACS) and vaso-occlusive pain crisis (VOC). Emerging evidence indicates acute elevations in circulating heme contributes to both ACS and VOC. Therefore, we tested the hypothesis that acute hemolytic crisis triggers AKI in SCD. We found that steady-state plasma A1M is raised significantly in SCD patients compared to controls (SS: 0.27±0.03, AA: 0.18±0.02 mg/ml p<0.05; n=12), and we determined that the molar A1M/Hx ratio is 14-fold higher in transgenic homozygous SCD (SS) mice compared to control (AA) littermates (n=6; p<0.05). Infusion of a non-lethal dose of purified heme (20 μmoles/kg) raised the concentration of heme in the liver but not in the kidneys of AA mice in 48 hours. On the contrary, the heme infusion over the same duration raised the total heme concentration in the kidneys but not in the liver of the SS mice (p<0.05). The SS mice but not the control AA mice developed severe renal tissue damage, defined by tubular atrophy and cast formation, loss of epithelial brush borders and apoptosis of the proximal tubular epithelium, within 48 hours of the heme infusion. Clinical detection of AKI is typically defined by elevation in serum creatinine and urinary albumin creatinine ratio (uACR). The acute elevation in circulating heme increased the plasma creatinine by 4-fold (p<0.001) with a 2-fold elevation in uACR (p<0.01) exclusively in the SS mice but not in control AA mice. Urinary kidney injury molecule-1 (KIM-1), which is a reliable predictor of AKI increased ~3-fold in the SS mice following the heme infusion, while no alteration of KIM-1 was evident in AA mice treated with the same challenge (n=6). To define the relationship between AKI in SCD and the hepatic and renal mechanisms of heme clearance, we measured plasma A1M and Hx, and urinary KIM-1 in a cohort of Ghanaian SCD patients aged 1-14 years (n=40). The molar ratio of A1M/Hx correlated positively with KIM-1 in this patient population (Pearson r=0.54; p=0.0003). Together, our data supports the conclusion that acute elevation of circulating heme triggers AKI in SCD. Clinically, the relative molar concentrations of plasma A1M and Hx can be developed as a biomarker to assess AKI risk in SCD patients. Finally, our data suggest acute infusions of Hx may protect SCD patients from AKI during episodes of hemolytic crisis. DisclosuresOfori-Acquah:NuvOx Pharma: Patents & Royalties.

Distinct Biomarkers Reflect Pathophysiological Differences of Sickle Cell Disease Sub-Phenotypes: Viscosity-Vaso-Occlusion Vs. Hemolysis-Endothelial Dysfunction

Background: Patients with Sickle Cell Disease (SCD) have variable phenotypes, with different severity of pain and other symptoms, including lung injury, stroke, leg ulcers, renal injury with proteinuria, osteonecrosis, and systemic and pulmonary hypertension. These phenotypes result from erythrocyte injury caused by HbS and its deoxygenation-induced polymerization. Recently, two distinct sub-phenotypes have been defined: a) Patients with the Viscosity-Vaso-Occlusion sub-phenotype (VVO) suffer mainly from vaso-occlusive pain crises with a relatively high hemoglobin concentration and b) patients classified as the Hemolysis-Endothelial Dysfunction sub-phenotype (HED) suffer from stroke and pulmonary hypertension with an elevated concentration of LDH (Kato et al. J Clin Invest 2017;127:750).Aim: In this context, we aimed to explore the correlation of key biomarkers with the two sub-phenotypes of the disease, namely: Placental growth factor (PlGF) a member of the vascular endothelial growth factor superfamily, which plays an important role in both inflammation and neo-angiogenesis; von Willebrand Factor antigen (vWF:antigen), a multimeric plasma glycoprotein secreted by the endothelium; Growth Differentiation Factor-15 (GDF-15), a member of the Transforming Growth Factor-beta superfamily, which expression is strongly upregulated in response to oxidative stress, inflammation and tissue injury, as well as in conditions related to ineffective erythropoiesis. We tested these biomarkers in patients with compound heterozygous SCD and beta-thalassemia (HbS/βthal).Patients and Methods: Ninety adult Caucasian patients with HbS/βthal were included in the study, while 20 apparently healthy individuals, of similar age and gender, served as controls. Patients with HbS/βthal were divided in two groups according to their LDH levels: High-LDH (LDH>270U/L) (HED-phenotype) group (42 patients) and the Normal-LDH (LDH<270U/L) (VVO-phenotype) group (48 patients). Along with hematologic and blood chemistry parameters determination, measurements of circulating levels of PlGF, vWF:antigen, GDF-15, hs-CRP, Cystatin C, hs-TnT and D-Dimers were performed in both groups of patients and controls using RUO and IVD immunoenzymatic techniques.Results: We found that patients with the HED-phenotype compared to the VVO-phenotype of the disease had lower Hb levels (p<0.001), higher Reticulocyte Production Index and higher serum bilirubin (p<0.001 and p=0.004, respectively), while there were no differences regarding Hb F levels between the two groups. PlGF levels were significantly elevated only in patients with the HED-phenotype (22.6±7.1pg/mL) compared to the controls (15.2±2.4pg/mL) (p<0.001) and patients with the VVO-phenotype (18.2±7.6pg/mL) (p=0.005). vWF:antigen concentrations were markedly elevated in both groups of patients compared to controls (186.4±81.7 and 157.8±73.4 vs 85.3±22.1IU/dL, p<0.001), with the increase of vWF:antigen levels to be more pronounced in patients with the HED-phenotype (p=0.008). Similarly, GDF-15 levels were also markedly elevated in both groups of patients compared to controls (2346.2±1295.6 and 1693.5±1398.3 vs 665.4±221.9pg/mL; p<0.001), with the increase of GDF-15 levels to be more pronounced in patients with the HED-phenotype (p=0.006). Regarding other parameters tested we found significant higher levels of D-Dimers in patients with the HED-phenotype (p<0.001) compared to patients with the VVO-phenotype, while no differences were found in parameters of inflammation and renal function.Conclusions: These findings demonstrate for the first time in the literature the correlation and involvement of PlGF, vWF:antigen and GDF-15 proteins in the pathophysiological mechanisms of the Hemolysis-Endothelial Dysfunction sub-phenotype of SCD in patients with the HbS/βthal genotype. Although, there is a degree of overlapping between the two sub-phenotypes of SCD, the differences in the specific biomarkers were significant. Thus, these markers along with the clinical profile could better identify the two subtypes of SCD patients and drive an innovative approach with the use of direct personalized therapies for each specific sub-phenotype by targeting the predominant mechanism in this multifactorial disorder. DisclosuresNo relevant conflicts of interest to declare.

Quantitative Sensory Testing in Children with Sickle Cell Disease

▪Background: Quantitative Sensory Testing (QST) is used in a variety of pain disorders such as arthritis and neuropathic pain syndromes, to characterize pain and predict prognosis and response to specific therapies. QST evaluates sensory function using a set of psychophysical procedures to measure subjective somatosensory responses to various physical stimuli. In Sickle Cell Disease (SCD) the painful vasoocclusive crisis is the most difficult management challenge and factors underlying the transition from acute to chronic pain and the wide interpatient variability in the pain experience are poorly understood. Optimal management for this complex pain syndrome remains unclear despite over three decades of basic and clinical research in the field.In this study, we aimed to duplicate results in the literature documenting altered QST thresholds in SCD and test the reproducibility of results over time to validate QST as a therapeutic endpoint for use in clinical trials evaluating novel preventative pain therapies.Objectives: Primary objectives were to test thermal thresholds in children with SCD versus African American controls and determine consistency over 6 months. Exploratory objectives were to correlate QST with clinical factors and select biomarkers and assess pressure pain thresholds (PPTs).Methods: Fifty-seven SCD and 60 control subjects aged 8-20 years underwent heat (wdt) and cold (cdt) detection and pain (hpt and cpt) threshold testing using a Medoc TSAII. A thermal composite score (TCS) was constructed by averaging the wdt, cdt, hpt, and cpt threshold responses from the testing instrument's starting temperature of 32°C. All participants were tested at baseline and 3 months; SCD subjects were additionally tested at 6 months and blood collected for future biomarker assessment. PPTs were performed using an SBMedic algometer. A single individual performed all QST. Descriptive statistics were computed and covariate-adjusted differences in thresholds within and between groups evaluated over time by mixed effects regression models with random intercepts and linear contrast tests. The intraclass correlation coefficient (ICC) from these models was used to assess within-subject consistency and Spearman's correlation was used to evaluate associations between thresholds and biomarkers. In addition, as a nested pseudo-intervention study, we analyzed a subset of “new” hydroxyurea (HU) users consisting of SCD subjects in whom HU was initiated within a few weeks prior to, or shortly after baseline testing. Significance was set at 0.05.Results: There were no significant differences in mean thermal or PPTs between SCD and control participants at baseline (Table 1) or month 3. QST measurements were consistent in SCD and control subjects for cpt, wdt, hpt and TCS (each ICC>0.55), but not necessarily for cdt (ICC = 0.37). Although generally internally consistent, there was some variability in mean thermal thresholds among SCD subjects over time, but the only significant difference between the 3 and 6 month means was for hpt (+0.67°C; 95% CI 1.15, 0.19). There was no association between mean thermal thresholds and chronic transfusion therapy. The group in whom HU was initiated shortly after baseline testing (n=6) demonstrated significant reductions in all thermal sensitivity and pain threshold measurements following HU initiation (Table 2).Conclusions: In contrast to Brandow's study (Brandow, American Journal Hematology, 2012), which documented thermal hypersensitivity in SCD, we have not demonstrated significant differences in thermal thresholds between SCD and control subjects. This could be due, in part, to differences in body sites used for testing. Results between second and third test sessions were highly consistent, indicating that reliability may improve once subjects have experience with QST. Subjects on whom HU was initiated shortly after baseline testing (new HU users) exhibited progressive and significant decreases in both cold and heat sensitivity from baseline to 6 months, suggesting that thermal testing is sensitive to effective therapy to prevent vasoocclusive pain. These findings are of critical importance to inform the planning of future SCD related clinical trials using QST as an endpoint in the evaluation of preventative pain therapies.Supported by National Institute of General Medical Sciences National Institutes of Health Award P20GM109021 and U54GM104941 [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Cold Pain Causes the Greatest Drop in Microvascular Blood Flow in Sickle Cell Disease Subjects and Normal Controls Exposed to Graded Thermal Stimuli

Background: Painful vaso-occlusive crises (VOC) are a frequent and debilitating complication of sickle cell disease (SCD). The exact mechanism of transition from steady state to VOC is not well understood although it is thought to occur due to progressive blockage of the microvasculature with rigid sickle shaped red blood cells. Thus, any trigger that decreases the microvascular blood flow increases the red blood cell transit time and can promote entrapment of sickled cells in the microvasculature and progression to VOC. Environmental factors such as exposure to cold wind and changes in weather are common triggers of VOC and have been shown to be associated with increased frequency of hospitalizations for pain in patients with SCD. There is limited experimental data in literature on the physiologic effects of these factors on peripheral perfusion in SCD.Objective: To study the effect of graded thermal stimuli on the peripheral microvascular blood flow in SCD.Methods: 17 SCD and 11 control (healthy or sickle trait) subjects aging 13 to 39 years were exposed to graded thermal stimuli via TSA- II Neurosensory analyzer thermode that was placed on the thenar eminence. Individual threshold temperatures for heat and cold detection, heat and cold pain were applied and changes in the microvascular blood flow were measured on the contralateral thumb using photo-plethysmography (PPG).The vasoconstriction response within the complex microvascular blood flow signal was detected using cross-correlation technique. In order to quantify the vasoconstriction response, we calculated mean blood flow (MBF) from the PPG amplitude during each of these stimuli and compared to the MBF at baseline.Results: Cross correlation analysis showed that cold pain caused significant vasoconstriction response in 75% of the subjects, followed by heat pain (57%), cold detection (39%) and heat detection (18%).There was a significant drop in the MBF during cold pain, heat pain and heat detection when compared to baseline MBF (p <0.001) where cold pain caused the greatest drop in MBF (Figure 1). SCD subjects trended towards greater percent change in MBF from baseline across all the threshold temperatures but there was no significant difference between the two groups. The average threshold temperatures were comparable across the two groups with no significant differences in thermal sensitivity detected between SCD and controls (Table 1).Conclusions: Exposure to graded thermal stimuli causes a progressive drop in microvascular blood flow with exposure to cold pain eliciting the strongest vasoconstriction response. The vasoconstriction occurred in the contralateral hand at an average of 10 seconds after the stimulus, suggesting a neurally mediated mechanism. Although there was no significant difference in vasoconstriction responses between SCD and controls, the drop in microvascular blood flow in sickle cell patients can increase the likelihood of entrapment of the sickled red blood cells in the microvasculature leading to vaso-occlusion. These findings are consistent with extensive reports in literature that exposure to extremes of cold weather is associated with a higher frequency of VOC. This suggests that neurally mediated decrease in the microvascular blood flow is likely an important factor in the pathophysiology behind cold exposure leading to VOC in SCD. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Pharmacokinetics of Hydroxyurea Therapy in African Children with Sickle Cell Anemia:Â Noharm Ancillary PK Study

▪Background. Hydroxyurea is an important disease-modifying therapy for sickle cell anemia (SCA). Through induction of fetal hemoglobin and other mechanisms, hydroxyurea reduces the number and severity of vaso-occlusive painful crises and episodes of acute chest syndrome, while also lowering the frequency of transfusions and hospitalizations, and even reducing mortality. In the US, hydroxyurea is recommended widely for both children and adults with SCA, although variation in treatment response is well-recognized. In sub-Saharan Africa, which has the greatest global burden of SCA, hydroxyurea is not used widely, due in part to a lack of data regarding its safety and efficacy. Pharmacokinetics (PK) analysis of hydroxyurea has also not been performed to date in Africa, but would provide important data regarding the variable responses to hydroxyurea treatment in low-resource settings.Objectives. There were three main goals of the NOHARM Ancillary PK Study: (1) to evaluate the feasibility of hydroxyurea sparse sampling PK collection and analysis in a low-resource clinical setting; (2) to determine the PK parameters for a cohort of young African children with SCA receiving hydroxyurea at an average oral daily dose of 20 mg/kg; and (3) to compare the PK parameters of African children collected and analyzed in the NOHARM trial with those of US children with SCA from the earlier HUSTLE study that included African-American children with SCA.Methods. NOHARM (NCT01976416) is a prospective, randomized, placebo-controlled, double-blinded phase 3 trial that is investigating the effects of hydroxyurea on the incidence and severity of malaria in very young children living in Uganda with SCA. After a year of blinded treatment, all participants were offered open-label hydroxyurea at an average oral daily dose of 20 ± 2.5 mg/kg, using either a 5-day or 7-day per week treatment regimen. After written informed consent for the ancillary PK study, hydroxyurea analysis was performed using a published sparse-sampling technique based on Bayesian estimation with collections pre-dose and then at T=15 minutes, 60 minutes, and 180 minutes. These four timed finger-prick sample collections over 3 hours used a volumetric absorptive microsampling device that collects exact 10 uL aliquots. All samples were stored at -80°C until analysis of hydroxyurea by LC-MS/MS (ug/mL) using a stable isotope internal standard, performed at Cincinnati Children's Hospital. Population PK analysis was performed using nonlinear mixed effect modeling (NONMEM 7.2). Allometric scaling was used to correct for differences in size.Results. A total of 100 children were enrolled in the NOHARM ancillary PK study (average age 3.8 ± 0.9 years; 55 males), and 98 of these children had three evaluable post-dose samples. Individual PK profiles were best described by a one-compartment model with first order absorption. The dose-normalized hydroxyurea concentrations in NOHARM were comparable between the 5-day and 7-day dosing regimens and similar to published HUSTLE data for 96 children, except for higher T=180 hydroxyurea concentrations and overall higher hydroxyurea exposure in the NOHARM cohort. The higher blood hydroxyurea concentrations were the result of reduced drug clearance in NOHARM, possibly influenced by younger age. Population PK parameter estimates of oral clearance in the NOHARM and HUSTLE cohorts were 9.8 L/h/70kg versus 12.9 L/h/70kg, respectively (p<0.01).Conclusions. The NOHARM PK study demonstrates the feasibility of collecting timed samples for analysis of hydroxyurea exposure, even in the low-resource setting of Uganda. Using a sparse sampling technique, young children with SCA had individual PK parameters calculated and a population PK model established. The oral drug clearance was lower in NOHARM than HUSTLE, with resulting higher hydroxyurea exposure, perhaps reflecting differences in age, nutrition, glomerular filtration, and genetic modifiers that warrant further investigation. DisclosuresWare:Agios: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Global Blood Therapeutics: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees.

Understanding Sleep in Pediatric Patients with Sickle Cell Disease Admitted for Vaso-Occlusive Pain Crisis through Objective Data

▪Introduction: Sickle cell disease (SCD) is an inherited red cell disorder that leads to sickling of red blood cells, anemia and vaso-occlusion. The most common reason for hospitalization and morbidity in children is pain due to vaso-occlusive crisis (VOC). Importantly, poor sleep quality can lead to increased pain the subsequent day and nocturnal pain leads to reduced deep sleep, both which can then modify pain sensitivity. Studies using sleep diaries have shown this cyclical relationship between sleep and pain. Frequent occurrences of restless sleep are therefore believed to contribute to an increased severity and intensity of pain episodes.There is very little data, however, looking at objective data such as vital signs to define sleep in patients with SCD admitted for VOC. We aimed to make comparisons between sleep hours and daytime hours for pain scores, patient controlled analgesia (PCA) usage and vital sign data in effort to better define and understand sleep in SCD.Methods: We performed a retrospective chart review for patients with SCD between January 1, 2014 and January 31, 2017. We included all patients admitted to the pediatric service, admitted for >48 hours for simple VOC, and placed on a PCA during admission. Patients with complications of acute chest syndrome, stroke or surgery were excluded. The primary aim was to describe sleep using objective data during sleep hours for patients with SCD admitted with pain. In this study, sleep was simply defined by time (12am-6am). Objective data included: PCA usage and nursing documentation such as vitals. Secondary aim was to examine potential differences during sleep to other 6-hour intervals of time for objective data. Differences within each patient were analyzed using Wilcoxon signed-rank test, a non-parametric statistical hypothesis test used for repeated measurements on the same sample to compare their population means.Results: We analyzed 50 patients that met criteria (46% female and 78% with HbSS). The median age was 14 (range 2-19) and median days of admission was 6 (range 4-11). Average length of stay was 6.3 days.We found each patient analyzed had extremely different patterns of pain scores, vitals, and PCA usage patterns at day and night during hospitalization for VOC. Mean and median values at night as compared to daytime hours for vitals (HR, systolic blood pressure - SBP, and diastolic blood pressure - DBP), PCA usage, and pain scores did not differ across all patients. In contrast, we did find most individual patient data revealed an increasing HR and blood pressure following admission, which then decreased prior to discharge.Interestingly, within patient analysis found significant differences between night hours and day hours. The number of PCA doses attempted and delivered were both significantly lower in sleep hours (z=-2.7, p=0.007 and z=-3.2, p=0.002). Vital signs indicated that SBP, DBP, and respiratory rate were all also significantly lower during sleep hours (see Table 1). We did not find differences between sleep and daytime hours for pain, HR and temperature.Conclusion: As expected, we found a large amount of variation across all patients for PCA usage and objective measures. Using a simple time definition of sleep, we also found no differences between night hours and daytime hours across all patients. However, while there were no statistically significant differences in vitals across the 50 patients between day and night, we found that the pain levels during their hospitalization to be highly individualized in nature.We found significant differences (at significance level of 0.05) between night hours and daytime hours within each patient. These included PCA usage, SBP, DBP, oxygen saturation, and respiratory rate. Although no differences were found in pain and HR, we did notice interesting trends during hospitalizations, including a gradual increase and eventual decrease in HR and blood pressure for many of the patients. This further highlights the need for clinical decisions and treatments to be tailored to individual patients based on their vitals and medication usage. We are optimistic to find significant differences in objective measures during sleep hours and aim to expand our analysis to determine potential prediction of sleep to subsequent day outcomes (pain and discharge) and definition of ‘good sleep’ by objective measures (such as low HR) using individual patient models. [Display omitted] DisclosuresJonassaint:Sicklesoft: Other: Owner. Shah:Novartis: Other: Speaker; Alexion: Other: Speaker.

Management of Infections in SCD Patients Treated with HU in the Framework of Escort HU

▪Background and methodThis study aims to investigate on infections in sickle cell patients treated with hydroxycarbamide (hydroxyurea, HU) for symptomatic sickle-cell syndromes enrolled in the multicentric prospective non-interventional cohort study ESCORT HU (European Sickle Cell Disease COhoRT - HydroxyUrea). The first patient was enrolled in ESCORT HU in January 2009, and by June 2017, 1844 patients were enrolled in the cohort.The increased susceptibility to infections of sickle-cell disease (SCD) affected patients is well known and likely results from several causes, including impaired splenic function, defects in complement activation, micronutrient deficiencies, and tissue ischemia. Splenectomy has been considered for a long time at high risk of early and late infections and of death in children (particularly aged 4 years or below) and adults with SCD. Although antibiotics and vaccines measures have been a huge step forward in preventive care, infections remain a major cause of morbidity and mortality in the developed countries and even more so in the developing world. Hydroxyurea (HU) is registered in Europe and in the US to prevent the recurrence of painful vaso-occlusive crises including acute chest syndrome. In Europe the marketing authorization for HU covers adults, adolescents and children older than 2 years suffering from symptomatic Sickle Cell Syndrome. HU is a well-tolerated fetal hemoglobin (Hb) inducer, with dose-dependent myelosuppression being the main short-term side-effect. Concerns have been raised about the possibility of HC as a predisposing factor for infections due to its myelosuppressive property.We present here a snapshot of infections collected in SCD patients enrolled in the prospective study cohort ESCORT HU. Children (≥ 2 years) and adults were enrolled from four European countries: France, Germany, Greece and Italy. Data are collected through an electronic Case Report Form (e-CRF). An analysis of these infections has been performed.ResultsFrom January 2009 until June 2017, 1841 patients (1016 adults and 822 children) have been enrolled in ESCORT HU. 746 infections were collected in the database in 437 patients (25.5%), of which 231 children and 206 adults. Hb Genotypes were SS (89.02%) or Sβ0 (4.35%) in 408/ 437 patients (93.3%) (see table 1).Features of the infections are reported in table 2. Bacterial infections were reported more frequently (42.4%), followed by viral infections including parvovirus B19 (26%). In 227 cases (31%), the type of infection was not specified. The spleen status was reported sporadically.As expected, infections were more frequent in children (52.4% in children vs 46.9% in adults), despite the fact that most of them received penicillin prophylaxis. Nevertheless the number of episodes of infection per patients was quite similar between adults and children: 350 infections were reported in 206 adults (1.70 per patients) and 386 in 231 children (1.67 per patient). 60% of patients (adults as well as children) experienced one episode of infection whereas 40% reported several infections. A small number of patients (33/431) had more than 3 infectious episodes. According to the investigators, infections were more severe in adults (15.14%) than in children (8.84%), but more frequently led to hospitalization in children than in adults (34.4% vs 24.4%). Antibiotics were widely prescribed for these episodes in 77.1% (adults) and 82.6% (children) of infections. No death due to infectious episode has been reported in the cohort.ConclusionsDespite an appropriate management of patients with sickle cell disease in Europe with antibiotic prophylaxis and recommendations for vaccinations (including routine vaccines and immunization against S. pneumoniae, N. meningitidis and H. influenzae), in Europe the incidence of infections remains high and is responsible for many hospitalizations. No increase of infectious risk has been reported in placebo-controlled clinical trials comparing HU and placebo in children (Baby HUG, MSH trial, SWitCH and TWitCH trials, Jain et al 2010) but comparisons with untreated SCD cohorts are required to better evaluate the incidence of infections in SCD patients treated with HU. [Display omitted] DisclosuresGalactéros:Addmedica: Membership on an entity's Board of Directors or advisory committees. De Montalembert:Addmedica: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Ribeil:Addmedica: Membership on an entity's Board of Directors or advisory committees; Vitalaire: Other: Non-financial support; Novartis: Other: Non-financial support. Kordes:Addmedica: Membership on an entity's Board of Directors or advisory committees. Grosse:Addmedica: Membership on an entity's Board of Directors or advisory committees. Brousse:Add Medica: Membership on an entity's Board of Directors or advisory committees. Oevermann:Addmedica: Membership on an entity's Board of Directors or advisory committees. Lobitz:Addmedica: Membership on an entity's Board of Directors or advisory committees.

Platelet Aggregation Dysfunction in Sickle Cell Disease

Acute systemic painful vaso-occlusive crisis (VOC) is the predominant clinical event requiring sickle cell disease (SCD) patients to seek emergency medical care. VOC often serves as an antecedent to acute chest syndrome (ACS), which is a type of acute lung injury and the leading cause of mortality in SCD. Current treatments for ACS are primarily supportive, and there is a critical need for rescue strategies to prevent the progression of VOC to ACS. ACS involves thrombosis across pulmonary artery branches, but the pathophysiology of thrombosis in SCD is largely unknown. Adenosine diphosphate (ADP) released from lysed erythrocytes can activate platelets by stimulating the purinergic receptors P2Y1 and P2Y12, suggesting that blocking these receptors may prevent VOC. Surprisingly, P2Y12 receptor blockade was unsuccessful in preventing VOC in a recent DOVE trial, justifying the need for better understanding of purinergic signaling in SCD.We used a novel intravital microscopy enabled experimental model of vaso-occlusive pulmonary thrombosis in transgenic SCD mice triggered by an intravenous (IV) administration of ADP. This method allows real time in vivo visualization of platelet aggregation in the lung of live SCD mice using multi-photon-excitation based quantitative fluorescence intravital lung microscopy (qFILM). We discovered that vaso-occlusive thrombosis involves entrapment of large circulating platelet aggregates in the bottle-necks located between the pulmonary arterioles and capillaries. Surprisingly, the dose of ADP that led to the pulmonary vaso-occlusive thrombosis in control mice, failed to induce vaso-occlusive thrombosis in SCD mice. Remarkably, further in vivo and in vitro platelet aggregation studies confirmed that SCD mouse platelets were indeed refractory to ADP but not collagen.Our preliminary findings suggest that therapeutic inhibition of P2Y1 and P2Y12 platelet receptors will not be a successful therapy in SCD. Our findings also warrant the need for further studies to understand the role of purinergic signaling in SCD DisclosuresNo relevant conflicts of interest to declare.