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Rationale: Intraerythrocytic polymerization of Hb S promotes hemolysis and vasoocclusive events in the microvasculature of patients with sickle cell disease (SCD). Although platelet–neutrophil aggr...

Acute lung injury, referred to as the acute chest syndrome, is a major cause of morbidity and mortality in patients with sickle cell disease (SCD), which often occurs in the setting of a vaso-occlusive painful crisis. P-selectin antibody therapy reduces hospitalization of patients with SCD by ∼50%, suggesting that an unknown P-selectin–independent mechanism promotes remaining vaso-occlusive events. In patients with SCD, intraerythrocytic polymerization of mutant hemoglobin promotes ischemia-reperfusion injury and hemolysis, which leads to the development of sterile inflammation. Using intravital microscopy in transgenic, humanized mice with SCD and in vitro studies with blood from patients with SCD, we reveal for the first time that the sterile inflammatory milieu in SCD promotes caspase-4/11–dependent activation of neutrophil–gasdermin D (GSDMD), which triggers P-selectin–independent shedding of neutrophil extracellular traps (NETs) in the liver. Remarkably, these NETs travel intravascularly from liver to lung, where they promote neutrophil-platelet aggregation and the development of acute lung injury. This study introduces a novel paradigm that liver-to-lung embolic translocation of NETs promotes pulmonary vascular vaso-occlusion and identifies a new GSDMD-mediated, P-selectin–independent mechanism of lung injury in SCD.

Introduction. Sickle cell disease (SCD) is caused by hemoglobin S polymerization leading to vaso-occlusion, small vessel disease, and reduced tissue oxygen saturation. Noninvasive assessments and screening of small vessel disease and decreased oxygen saturation in select organs, like the brain, are poorly developed. Cerebral autoregulation, a mechanism that ensures cerebral blood flow (CBF) remains constant with changes in mean arterial pressure, is impaired with and is a marker of small vessel disease. Frequency-domain near-infrared spectroscopy (FD-NIRS) can be used as a noninvasive technique to measure cerebral autoregulation by recording cerebral microvascular changes in hemoglobin concentration that are related to cerebral blood flow. Transfer function analysis is a method to noninvasively quantify the relationship between measured physiologic signals, and can be used to estimate cerebral autoregulation. In this study, we hypothesized patients with SCD have impaired cerebral autoregulation and reduced tissue oxygen saturation, which we evaluated via FD-NIRS with transfer function analysis, as compared to healthy controls. Methods. All patients provided informed consent under an IRB protocol approved by the University of Pittsburgh and Carnegie Mellon University. Patients with SCD (n = 11) and healthy, race-matched controls (n =14) were enrolled. All participants sat upright in a chair with probes of a commercial multi-distance FD-NIRS apparatus applied to their forehead in a dark room (Fig. 1A). We used paced breathing to dynamically induce controlled changes in the microvascular tone at 0.1, 0.125, and 0.1667 Hz, spanning normal to more challenging breathing rates, with a visual metronome guiding each paced breathing session (Fig.1.B). To validate each breathing pace, we recorded the participants' respiratory rate via a belt placed around their chest, and their mean arterial pressure through a finger and arm cuff attached to a beat-to-beat finger plethysmographer (Fig.1.A). We used the modified Beer-Lambert law to calculate changes in oxy-hemoglobin (O), deoxy-hemoglobin (D), and total hemoglobin (T) concentrations. Data was bandpass-filtered around the paced breathing frequencies, and the Hilbert transform was applied to calculate the amplitude ratios, |D|/|O|, and phase latency, Arg(D)−Arg(O). The tissue oxygen saturation of hemoglobin, StO2 (mean ±SD), was computed based on an average baseline of two minutes. Results. Arg(D)−Arg(O) and |D|/|O| are shown for controls and patients at all paced breathing frequencies (Fig.1C-D) from the largest source-detector distance. A non-zero phase lag between D and O within both groups indicates influences of cerebral blood volume (CBV) and CBF changes. The phase differences of D and O are therefore related to cerebral autoregulation. We found a greater phase difference (mean range: -320° to -340°) in patients with SCD compared to controls (mean range: -200° to -240°), indicating the presence of vascular obstruction and impaired cerebral autoregulation. Higher amplitude ratios |D|/|O| may indicate slower blood transmit times. We found that patients with SCD had increased amplitude ratios for hemodynamic oscillations at lower paced breathing frequencies compared to healthy controls (Fig. 1D). Finally, we found a significantly reduced cerebral tissue oxygen saturation in patients with SCD (63.1 % ± 7.8 %) in comparison with controls (65.9 % ± 4.9 %, p Discussion. We provide preliminary evidence that FD-NIRS can be used to assess cerebral autoregulation in SCD. Our results confirm and extend those of a prior study by Reinhard et al, 2003, by using the phase latency, Arg(D)−Arg(O), and the amplitude ratios, |D|/|O|, to assess autoregulation. In addition, we detected a noticeable reduction in oxygen saturation in patients with SCD as compared to healthy controls. Further studies may extend our findings to additional frequencies in a larger sample size to more comprehensively assess the impact of mean arterial pressure on autoregulation. Noninvasive measurement of cerebral autoregulation and brain oxygenation in SCD by FD-NIRS may represent a novel biomarker of cerebral vasculopathy in SCD and may help monitor changes in cerebral oxygenation, particularly during treatment with drugs that modulate the hemoglobin oxygen affinity, such as the recently FDA-approved voxelotor. Figure 1 Disclosures No relevant conflicts of interest to declare.

Author(s): Jonassaint, Charles R; Kang, Chaeryon; Abrams, Daniel M; Li, Jingyi J; Mao, Jason; Jia, Yimeng; Long, Qi; Sanger, Maureen; Jonassaint, Jude C; De Castro, Laura; Shah, Nirmish

Background. Sickle cell disease (SCD) remains the most common genetic hematologic disorder, with a disproportionally high incidence and prevalence in African countries. It is associated with an increased risk of maternal and infant morbidity and mortality compared to the general population. As more young adults living with SCD reach healthier reproductive ages, it is imperative that there is open communication between providers and patients regarding reproductive health, maternal risks associated with childbirth, and the risk of having children who inherit SCD. It is also important that providers understand reproductive health knowledge and perceptions within this population and methods to improve family planning education. We performed a survey-based study aimed to quantify current knowledge and perceptions regarding reproductive health choices, family planning and genetic counseling-presented in another abstract-. Here we present a sub-analysis, aiming to quantify differences, based on the participants region, concerning reproductive health knowledge and perceptions amongst adults with SCD. Methodology. All study participants were at least 18 years old and have a SCD diagnosis. They were asked to complete an anonymous survey instrument consisting of 43 questions, administered using Qualtrics Survey Software. The survey was comprised of questions about participants' demographics, knowledge base and perceptions about reproductive health and genetic counseling. Participants were recruited using convenience sampling in two ways. The first was through reaching sickle cell patient groups online via social media through Facebook and Twitter. The second was at an adult SCD clinic where participants were invited to complete the survey in clinic or afterwards. 152 participants accessed the survey, and 105 that completed more than 90% of the survey were included in the primary analysis. Further analysis was done to compare data from 70 participants who did not migrate from their reported birth country. Comparison groups were generated using reported birth country and current location. Forty-seven participants from North American (U.S. and Canada) and 23 from African Countries (Nigeria, Zambia, Kenya, and South Africa) were included in this analysis. Results. Demographics: Among the 70 analyzed, 47 (67.1%) respondents were from North America (N.A.) and 23 (32.9%) were from African Countries (A.C). Knowledge: People from both N.A. and A.C. agreed that women with SCD were at higher risk of pregnancy complications (91.5% and 87%, respectively), and understood the chance of having a child with SCD if both parents have the trait (78.7% and 78.3%). Perception: Participants from N.A. reported receiving most of their information about family planning, partner screening, and reproductive health from healthcare providers more frequently than participants from A.C. (67% vs 39%). More participants from A.C, 21 (91.3%), agreed that having children in the future was important to them, while 21 (45%) of the participants from N.A agreed to this. Participants from A.C. and N.A. equally agreed that is it important to know if their partner has the sickle cell trait (SCT) (95.7% and 93.6%). However, more participants from A.C., 21 (91.3%), reported having any discussion with their partner about being screened for the SCT, versus 29 (61.8%) from N.A. Participants from N.A. and A.C. were equally knowledgeable about what a genetic counselor is (72% v. 70%, respectively), and utilized genetic counseling at low rates (37% v. 39.1%, respectively). Most participants reported that they do not know how to get in contact with a genetic counselor in their community, 31 (66%) from N.A. and 18 (78.3%) from A.C. Many have not been referred to a genetic counselor in the past by a healthcare provider, 39 (83%) from N.A. and 20 (87%) from A.C. Conclusion. Our study suggests that there are differences in reproductive health and genetic counseling knowledge and perceptions when comparing SCD participants living in two world regions. Independently of regional differences, many people with SCD appear to lack key reproductive health and genetic counseling knowledge and resources. We believe there is a need for improved communication between providers and patients in family planning education, as well as further studies to address access to reproductive health and genetic counseling resources. Disclosures Nunez: American Board of Genetic Counseling: Membership on an entity's Board of Directors or advisory committees; Foundation for Women and Girls with Blood Disorders: Membership on an entity's Board of Directors or advisory committees. De Castro:Novartis: Membership on an entity's Board of Directors or advisory committees; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy.

Purpose: Therapeutic erythrocytapheresis, known also as automated red blood cell exchange transfusion (RBC exchange), is standard of care for patients with sickle cell disease (SCD) presenting with severe complications and for primary prevention of CNS events in those with increased risks. There is limited data on the framework and experience of performing this procedure in a large adult population and even less on challenges and outcomes. Here, we present demographic and statistical data highlighting characteristics of the chronic exchange transfusion program at a comprehensive and multidisciplinary adult SCD center. Methods: Data were collected from adults (18 years and older) with a diagnosis of SCD that were receiving blood transfusions in the outpatient treatment facility from September 1st, 2017 to August 31st, 2018. Data gathered included age, gender, procedure indications and frequency, hemoglobin S percentage (Hb S%), target pre and post Hb S%, ferritin trend, and type of intravascular device used for procedure. Three hundred and forty-four (344) episodes of transfusions, were performed in 52 unique patients, corresponding to 21 % of the unique patients treated at our center during that period. Two hundred and sixty-two (262) of the transfusion procedures were RBC exchanges. These RBC exchanges were performed in 35 (14%) of the unique patients. Since a sizable proportion of our patients received pain treatment during RBC exchange, we explored how the pain visual analog score (VAS) changed while undergoing pain treatment during RBC exchange and in relation to multiple opiate doses. Descriptive and basic statistical analysis were performed on the data obtained for those receiving RBC exchange alone or in combination with pain management. Results: Thirty-five unique patients underwent RBC exchange during the study period. All but two (2) patients had the procedure via an implanted dual lumen Vortex port, one (1) by an arteriovenous fistula and one (1) via his native veins. Most of the RBC exchanges were performed as part of a chronic transfusion program for one of the following indications: primary prevention of stroke; secondary prevention of stroke; recurrent acute chest syndrome (ACS); intractable pain; presence of multiple comorbidities including pulmonary hypertension, chronic kidney disease and intractable leg ulcers. One patient received RBC exchange in preparation for bone marrow transplant. The three (3) other patients that only received one (1) RBC exchange during that period were previously on exchange transfusion but discontinued the procedure due lack of venous access or benefit from the procedure. The frequency of RBC exchange varied depending of the patient's specific indication for the procedure and the ability to reach a target Hb S% goal of < 30%. The RBC exchange occurred in patients as frequently as every four (4) to seven (7) weeks. The number of transfusions per patient/year ranged from one (1) to thirteen (13) with a mean and median of eight (8). Number of RBC units used per procedure ranged from four (4) to thirteen (13) with a mean of eight (8.6), with each unique patient usually receiving the same amount. Twenty-three (66%) unique patients received, at least once, acute pain treatment - oral, intravenous, or a combination during RBC exchange. The mean change of VAS between presentation and end of the pain management period was 1.9 (p Conclusion: Our data shows that therapeutic RBC exchange are performed on a regular basis on a small but significant subgroup of patients living with SCD and treated at a comprehensive SCD center. Intravenous opioid management of acute pain can be effectively performed while receiving RBC exchange which provides patients with multimodal interventions in the outpatient setting. The relationship between change in VAS and concurrent transfusion deserves further investigation. Although there are standard indications for RBC exchange, we and others use it for indications for which there is low level of evidence or no evidence of efficacy (i.e. leg ulcers). We found that there great variability on the frequency of the procedure, total number of RBC exchanges per year and number of RBC units used. Prospective studies are needed to better understand the short and long-term effects, complications and outcomes of chronic RBC exchange in adults living with SCD. Figure Disclosures Kato: Bayer: Research Funding; Novartis, Global Blood Therapeutics: Consultancy, Research Funding. De Castro:Pfizer: Consultancy; Novartis: Membership on an entity's Board of Directors or advisory committees; Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Background. With improvements in early diagnosis and health care for those with sickle cell disease (SCD), many affected are living longer and having families of their own. Despite medical advancements in this field, pregnancy in this population is still associated with an increased risk for maternal and infant morbidity and mortality compared to the general population. Due to increased maternal risks associated with childbirth and the risk of having children who inherit the disease, communication between providers and patients regarding reproductive health is imperative. However, providers' understanding of reproductive health knowledge and perceptions within this population and methods to improve family planning education which can result in autonomous and informed decisions amongst adults with SCD is still limited. Our study aims to quantify current knowledge and perceptions in adults with SCD regarding reproductive health choices and genetic counseling. Methodology. Study participants had to be at least 18 years old and have a diagnosis of SCD. Participants were asked to complete an anonymous survey instrument consisting of 43 questions that was administered using Qualtrics Survey Software. The survey was comprised of questions about participants' demographics, knowledge base and perceptions about reproductive health and genetic counseling. Participants were recruited using convenience sampling in two ways: 1) through online social media via Facebook and Twitter sickle cell patient groups; 2) patients at an adult sickle cell clinic were invited to complete the survey in clinic or afterwards. A total of 152 participants accessed the survey, and of those, 105 that completed more than 90% of the survey, were used in the analysis. Results. Demographics: Participants were between 18-57 years of age, 85 females (81.0%), 19 males (18.1%), and 1 transgender (1.0%). Among 99 people who disclosed their place of birth, 49 (49.5%) were from North America (U.S. and Canada), 38 (38.4%) were from African countries, and 12 (12.1%) were from other parts of the world. Fifty-six (53.5%) reported having no children and 49 (46.7%) had a minimum of one child. Knowledge: Ninety-three (88.6%) agreed that women with SCD are at a higher risk of pregnancy complications and 82 (78.1%) understood the chance of having a child with SCD if both parents have the trait. However, one-third (33.3%) did not know that women with hemoglobin SS will always pass down the trait to their children, and 90 (85.7%) were unaware of contraceptive methods available that have been shown to reduce the risk of sickle cell crisis. Perception: Sixty-four (61.5%) participants agreed that having children in the future was very important to them. Majority, 96 (91.4%), also agree that they want to avoid having a child with SCD. Fifty-eight (55.2%) indicated they received a majority of their information about partner screening, and reproductive health from a healthcare provider while 47 (44.8%) reported receiving most of their information from other sources (school, family members, friends, and independent research). Fifty-nine (59.6%) out of the 89 participants who responded agreed that they wished they had more conversations about partner screening and reproductive health with their hematologist. Seventy-five (72.1%) respondents reported that they know what a genetic counselor is and 40 (39.2%) reported having formal genetic counseling in the past. Among those who know what a genetic counselor was, only 30 (40.0%) actually knew how to get in contact with a genetic counselor in their community. Of those that have had formal genetic counseling in the past 15 (37.5%) reported having formal genetic counseling by a genetic counselor, while 25 (62.5%) had some form of counseling by another healthcare provider (i.e. hematologist, gynecologist, PCP, other); and 24 (60%) had one child or more. Conclusion. Our study suggests that many individuals with SCD may still lack important reproductive health and genetic counseling knowledge and resources. We believe that improved communication between healthcare providers and SCD patients is wanted and needed by this population to prevent pregnancy-related complications and improve outcomes. Future studies should focus on genetic counseling access, and reproductive health and family planning education to promote ongoing discussions and increase knowledge of both providers and individuals with SCD. Disclosures Nunez: American Board of Genetic Counseling: Membership on an entity's Board of Directors or advisory committees; Foundation for Women and Girls with Blood Disorders: Membership on an entity's Board of Directors or advisory committees. De Castro:Global Blood Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy.

Sickle red blood cells (SSRBCs) adhere to both endothelial cells (ECs) and the extracellular matrix. Epinephrine elevates cyclic adenosine monophosphate in SSRBCs and increases adhesion of SSRBCs to ECs in a β‐adrenergic receptor and protein kinase A‐dependent manner. Studies in vitro as well as in vivo have suggested that adrenergic stimuli like epinephrine may contribute to vaso‐occlusion associated with physiologic stress. We conducted both animal studies and a Phase I dose‐escalation study in sickle cell disease (SCD) patients to investigate whether systemically administered propranolol inhibits SSRBC adhesion and to document the safety of propranolol in SCD. Systemically administered propranolol prevented SSRBC adhesion and associated vaso‐occlusion in a mouse model. In patients receiving a single oral dose of 10, 20, or 40 mg propranolol, SSRBC adhesion to ECs was studied before and after propranolol, with and without stimulation with epinephrine. Propranolol administration significantly reduced epinephrine‐stimulated SSRBC adhesion in a dose dependent manner (p = 0.03), with maximum inhibition achieved at 40 mg. Adverse events were not severe, did not show dose dependence, and were likely unrelated to drug. No significant heart rate changes occurred. These results imply that β‐blockers may have a role as antiadhesive therapy for SCD. Clin Trans Sci 2012; Volume 5: 437–444

Introduction: Pediatric Blood and Marrow Transplant (PBMT) is an intense treatment fraught with significant symptom burden for patients. Mobile health (mHealth) and wearable technologies provide patient generated health data which may enhance understanding of complex symptom patterns, trajectories, and interactions. Improved monitoring and understanding may result in the development of precision symptom management strategies, yet limited research exists for integrating these technologies into acute patient care. Methods: We conducted a pilot study to determine the feasibility of integrating mHealth technology into symptom management for PBMT patients. Patient data includes: 1) an Apple Watch to collect objective data (heart rate, daily step count), and 2) a self-developed app to collect subjective symptom data. Patients were followed for up to 120 days, which could include both inpatient and outpatient days. Patients were asked to record within the mobile app daily and keep the wearable device (Apple Watch) on throughout each day and night with removal only to charge. The number of days the mobile app and wearable device were used by each patient was assessed to determine compliance to study protocol. In addition, transplant, engraftment, hospital discharge, and study discharge dates were used as 4 discrete time points to determine study feasibility. For transplant and engraftment, the patient must have recorded in the app within a week of the event to be considered an interaction to account for increased fatigue and duress during these times. On completion of the study, patients also completed feasibility questionnaires and interviews. Results: Twelve patients were approached and 10 patients enrolled in the study. Three patients withdrew early from the study citing the devices were to difficult to manage with their health status. Of the remaining 7 patients included in analysis, one went to the ICU on day 40. On average, the 7 patients charted data in the app for an average of 46.10% of their days involved in the study and the most commonly reported symptoms included pain (52%), fatigue (13%), fever (9%), and rash (9%). Patients wore the wearable device during 40.29% of their total days in the study. From their compiled 551 days in the study, this equates to 254 total days of app interaction and 222 days of using the wearable (Figure 1). Four patients completed all four discrete time points. Two patients are currently in the study (one has completed hospital discharge but not study discharge, and one has completed transplant and engraftment). For app interaction during discrete time points, the patients interacted with the mobile app 17 of 23 times. Furthermore, there was a 100% app usage during transplant and engraftment periods. When evaluating the app data jointly with the wearable data, it was determined that 71% of patients were interactive with the app and used the wearable device during transplant and 57% of patients were compliant with both the app and wearable device during engraftment. Interview data of all patients at study completion (n=4) found the devices easy to use, useful, and helpful, with comments: "I liked it because it was easy to keep track of how I felt. I liked looking back and comparing how I felt as the days went by". However, when they felt poorly, they didn't use them as frequently, "sometimes I just didn't feel good and didn't want to use them". Conclusion: There has been significant patient interest in participating in the study with the majority of patients approached also enrolling in the study. Study patients had variable compliance, which on average was nearly every other day. Limitations to app included patient fatigue and forgetfulness. Pain, itching, and battery life limited wearable compliance. Our findings suggest it is feasible to obtain data from mobile devices, although need to account for gaps in data due to significant symptoms and complications. Daily data collection may be difficult for acutely ill PBMT patients, however, combining both active and passive data collection measures may improve symptom cluster understanding and result in better symptom management interventions and strategies. Disclosures Shah: Novartis: Research Funding, Speakers Bureau.

Up to 30% of adult patients with sickle cell disease (SCD) will develop pulmonary hypertension (pHTN), a complication associated with significant morbidity and mortality. To identify genetic factors that contribute to risk for pHTN in SCD, we performed association analysis with 297 single nucleotide polymorphisms (SNPs) in 49 candidate genes in patients with sickle cell anemia (Hb SS) who had been screened for pHTN by echocardiography (n = 111). Evidence of association was primarily identified for genes in the TGFβ superfamily, including activin A receptor, type II–like 1 (ACVRL1), bone morphogenetic protein receptor 2 (BMPR2), and bone morphogenetic protein 6 (BMP6). The association of pHTN with ACVRL1 and BMPR2 corroborates the previous association of these genes with primary pHTN. Moreover, genes in the TGFβ pathway have been independently implicated in risk for several sickle cell complications, suggesting that this gene pathway is important in overall sickle cell pathophysiology. Genetic variation in the β-1 adrenergic receptor (ADRB1) was also associated with pHTN in our dataset. A multiple regression model, which included age and baseline hemoglobin as covariates, retained SNPs in ACVRL1, BMP6, and ADRB1 as independently contributing to pHTN risk. These findings may offer new promise for identifying patients at risk for pHTN, developing new therapeutic targets, and reducing the occurrence of this life-threatening SCD complication.

Sickle red cell (SS RBC) adhesion is thought to contribute to sickle cell disease (SCD) pathophysiology. SS RBC adhesion to laminin increases in response to adrenaline stimulation of beta(2)-adrenergic receptors (beta(2)ARs) and adenylate cyclase (ADCY6), and previous evidence suggests such activation occurs in vivo. We explored whether polymorphisms of the beta(2)AR and ADCY6 genes (ADRB2 and ADCY6, respectively) affect RBC adhesion to laminin. We found that the beta(2)AR arg(16)-->gly substitution and two non-coding ADCY6 polymorphisms were associated with elevated adhesion. We postulate that ADRB2 and ADCY6 polymorphisms may influence SCD severity through the mechanism of RBC adhesion.

Priapism, a painful and prolonged erection, has been reported to occur in 30–45% of male patients with sickle cell disease (SCD). However, little is known about the pathological processes and genetic risk factors that contribute to the occurrence of priapism. The identification of genetic variables that are associated with priapism may therefore help define both critical pathophysiologic mechanisms not otherwise apparent, as well as patients at increased risk. We examined genetic variation in our sample of 199 unrelated, adult (>18 years), male patients with Hb SS and Hb Sβ0-thalassemia, 83 (42%) of whom reported a history of priapism. Candidate genes for association with priapism were identified based on their involvement in adhesion, coagulation, inflammation, and cell signaling. Additionally, we examined genes involved in NO biology (NOS2, NOS3, SOD1, SLC4A1). Finally, we also examined polymorphisms in the KLOTHO gene, which has previously been associated with priapism. We examined a total of 389 SNPs in 48 candidate genes. Except for the gene encoding the β2 adrenergic receptor, SNP genotyping was performed by TaqMan, using Assays-on-Demand or Assays-by-Design genotyping products (Applied Biosystems). Allele tests were used to detect genetic associations with priapism. Strong evidence of association was found for SNP rs7526590 in the transforming growth factor-β receptor, type III (TGFBR3) gene (p=.00058), SNP rs10244884 in the aquaporin (AQP1) gene (p=.00068), and SNP rs3768780 in the integrin αV (ITGAV) gene (p=0.00090). A second ITGAV SNP (rs3768778), in linkage disequilibrium (r2=.59) with the first, also showed association with priapism (p=.00888). The A1 subunit of coagulation factor XIII (F13A1) had four SNPs (hcv1860621, rs1032045, rs1674074, rs381061) with p-values less than 0.010 (p-values = 0.00156, 0.00415, 0.00648, and 0.00712, respectively). The linkage disequilibrium among these F13A1 SNPs is negligible (r2

Introduction: Cognitive impairment is a serious complication of sickle cell disease (SCD) that affects both children and adults. While it is known to adversely affect school performance and overall development in children, its functional impact in adults is not known. Importantly, poor disease self-management and adherence to hydroxyurea are major problems in SCD and are likely to be impacted by cognitive impairment. In the study presented herein, we hypothesized that lower cognitive function in areas of memory and executive function would be associated with worse adherence to hydroxyurea therapy in adults with SCD. Methods: We designed a cross-sectional study of patients with SCD from the University of Pittsburgh Medical Center Adult Sickle Cell Clinic. Patients who underwent neurocognitive testing between 2011 and 2016 as part of an ongoing neurocognitive study and whose adherence to hydroxyurea therapy has been monitored by a dedicated clinical nurse were included. Performance on four neurocognitive tests of verbal learning, memory and executive function - the Hopkins Verbal Learning Test-Revised (HVLT-R) retention, HVLT-R total recall, Delis-Kaplan Executive Function System Trail Making Condition 4 vs. 5, and Color Word Condition 3 - were selected a priori as explanatory variables. We explored the association between the four test scores and the most recent adherence data at the time of the study. Adherence measures included the laboratory biomarkers red blood cell corpuscular volume (MCV) and percentage of fetal hemoglobin (HbF) - whose values are expected to rise with hydroxyurea therapy - and the patient's self-report of adherence categorized as "good" (≥ 80% adherence) or "less-than-good" (< 80% adherence). Differences between groups were compared by 2-sided t-test or chi-square tests where indicated. Results: Study participants (n=48) had a mean age of 35.2 years (range 22-60) and were predominantly female (58.3%). Fifty-two percent had higher than high-school level education, and 72.9% had a HbSS or HbS/β0 thalassemia phenotype. There were no statistically significant differences between the "good" adherence and "less-than-good" adherence groups with regard to these baseline characteristics. As expected, the mean MCV, an objective marker of hydroxyurea adherence, was significantly higher in the "good" adherence group (102.3 ± 15.0 vs. 89.1 ± 11.5 fL, p=0.005), corroborating the patients' self-report. We observed a statistically significant positive correlation between the HVLT-R retention score and MCV that remained significant after adjustment for demographic characteristics (Pearson r=0.309, p=0.033) and a similar correlation with HVLT-R total recall (Pearson r=0.269, p=0.065). A similar trend between HVLT-R retention and HbF percentage was observed (Pearson r=0.260, p=0.081). The mean HVLT-R retention and HVLT-R total recall scores were higher in the "good" adherence group (80.1% ± 27% vs. 64.9% ± 33%, and 41.9% ± 14% vs.34.1% ± 13%, respectively), although the difference between the two groups did not reach statistical significance. There was no significant difference in performance on the Trail Making Condition 4 vs. 5 or the Color Word Condition 3 tests. Discussion: Our study suggests that adherence to hydroxyurea therapy in adult patients with SCD might be affected by impairment in episodic memory, as measured in our study through HVLT-R retention, with qualitatively similar patterns by HVLT-R total recall. Although more extensive investigation is warranted, these results suggest that a brief, targeted neurocognitive assessment should be considered as a component of any strategy aimed at improving adherence to hydroxyurea and other therapeutic interventions in patients with SCD. Disclosures Jonassaint: Sicklesoft: Other: officer of Sicklesoft. Kato:Mast Therapeutics: Consultancy; Bayer: Research Funding.

Introduction: Sickle cell disease (SCD) is a chronic illness associated with frequent medical complications and hospitalizations. Importantly, approximately ninety percent of hospitalizations are for pain events, and 30-day re-hospitalization rates are alarmingly high. Factors influencing the high rates are poorly understood; however, close follow-up and continued titration of pain medications have been shown to decrease re-hospitalization rates. Mobile technology has become an increasingly integral part of health care management. We have recently developed SMART (Sickle cell Mobile Application to Record symptoms via Technology) to assist with documentation of pain and interventions. We propose using this current mobile technology as a tool for both patients and providers to monitorand manage symptoms and health interventions. We hypothesize that this technology will lead to an increase in follow up, a significant decrease in readmission rates, as well as overall disease burden. Methods: We randomized SCD patients who presented to the day hospital for pain control to either standard of care or an iPad with SMART. Patients were enrolled upon discharge from the day hospital and expected to return for a visit at 12 and 30 days after hospitalization. They were followed for a total of 30 days to determine re-utilization rates. Re-utilization was defined as a return for pain management to the day hospital, emergency department (ED), or admission to the hospital. Patients with more than 10 acute care visits in the past 12 months or who were on chronic transfusions were excluded. Standard of care (SOC) included a review of instructions, discharge medications, upcoming appointments and phone numbers for contact. SMART was programmed with the patient's scheduled medications, upcoming appointments and allowed patients and the medical team to communicate via text. Patients were provided a reminder through the app to record their symptoms and interventions (medication or non-pharmacologic) twice daily. The medical team reviewed symptoms recorded by patients at least once daily and texted patients who were indicating significant changes in pain. Results: We enrolled 19 SCD patients and randomized 10 to SMART and 9 to SOC (52% male, mean age 31.2 ± 6.6 years). Patients using SMART were more likely to return for their scheduled follow-up visit at 12 days (80% vs. 33%) and 30 days (80% vs. 44%). Thirty-day re-utilization of care was lower for patients using SMART compared to patients receiving SOC (20% vs. 77%, see Table 1). Furthermore, re-utilization for patients using SMART was limited to 2 visits to the day hospital, which occurred 10 and 24 days post discharge. This is in contrast to 8 visits to the ED and 3 to the day hospital among 7 of the 9 patients receiving SOC, with re-utilization occurring a median of 2 days post discharge (range 1-23). Of the patients using SMART, one patient did not make any entries into the application, while the other 9 patients made almost 1 entry per day for 30 days (median entries 28.9, range 5-107). The medical team texted all patients with SMART (range 2-19 times over 30 days); 60% returned texts to the medical team (range 2-11 times over 30 days). Interestingly, the two patients with SMART who returned within 30 days to the day hospital for further pain treatment had not returned any texts to the medical team. Conclusion: Our pilot study used an innovative mobile application to improve care for SCD patients who were discharged following day hospital treatment for pain crisis. Patients using SMART were more likely to return for their scheduled follow-up appointments. In addition, 30-day re-utilization of care was much higher for patients receiving SOC than SMART and was similar to previously observed institutional re-utilization rates. In contrast, only 20% of patients given SMART returned for re-utilization of care within 30 days, and they were the patients who did not use the texts for communication. We plan to expand the use of SMART to improve decision-making by medical providers through access to symptoms recorded by patients following discharge and, ultimately, to decrease re-utilization of care. Disclosures Jonassaint: Sicklesoft: Other: officer of Sicklesoft. Shah:Novartis: Speakers Bureau.

Sickle cell disease (SCD) is the most common inherited hemoglobinopathy worldwide. Our previous results indicate that the reduced oxidative stress capacity of sickle erythrocytes may be caused by decreased expression of NRF2 (Nuclear factor (erythroid-derived 2)-like 2), an oxidative stress regulator. We found that activation of NRF2 with sulforaphane (SFN) in erythroid progenitors significantly increased the expression of NRF2 targets HMOX1, NQO1, and HBG1 (subunit of fetal hemoglobin) in a dose-dependent manner. Therefore, we hypothesized that NRF2 activation with SFN may offer therapeutic benefits for SCD patients by restoring oxidative capacity and increasing fetal hemoglobin concentration. To test this hypothesis, we performed a Phase 1, open-label, dose-escalation study of SFN, contained in a broccoli sprout homogenate (BSH) that naturally contains SFN, in adults with SCD. The primary and secondary study endpoints were safety and physiological response to NRF2 activation, respectively. We found that BSH was well tolerated, and the few adverse events that occurred during the trial were not likely related to BSH consumption. We observed an increase in the mean relative whole blood mRNA levels for the NRF2 target HMOX1 (p = 0.02) on the last day of BSH treatment, compared to pre-treatment. We also observed a trend toward increased mean relative mRNA levels of the NRF2 target HBG1 (p = 0.10) from baseline to end of treatment, but without significant changes in HbF protein. We conclude that BSH, in the provided doses, is safe in stable SCD patients and may induce changes in gene expression levels. We therefore propose investigation of more potent NRF2 inducers, which may elicit more robust physiological changes and offer clinical benefits to SCD patients. Trial registration: ClinicalTrials.gov NCT01715480.

Sickle cell disease (SCD) is a chronic disease for which complications include acute chest syndrome, painful vaso-occlusive crisis, and stroke. There has been increased use of chronic transfusions to prevent and treat complications associated with SCD (Blinder, 2013), however, the result is iron overload for which complications include liver failure, heart failure and death. (Niederau, 1996; Brittenham, 1994; Harmatz, 2000) To effectively prevent and treat iron overload, chelation with oral medications must be taken correctly (Kwiatkowski, 2004). Depending on the method used to measure medication compliance, adherence varies between 43% and 76%. (Raphael et al., 2009; Alvarez et al., 2009) The variability underscores the need to better investigate and improve adherence. The success of treatment plans requires collaboration among patients, families and providers to take prescribed medications. We sought to implement mobile technology to facilitate this collaboration, while at the same time providing education and engaging both patients and parents. We performed a feasibility study using an original mobile application to assess adherence with oral chelation therapy and provide a platform for education modules. Eligible patients had a diagnosis of SCD or thalassemia with a history of iron overload, were ≥5 years of age and had access to a phone or other mobile device with the iOS operating system. Devices were provided for use if needed. During the first 30 days of the study, participants were asked to self-record up to a 5-minute video of daily medication administration. They performed this with our mobile application, which we could review and track. On day 2-3 of participation, they received a generic welcome message via the application. On day 7 of participation, they received a personalized message of encouragement for consistent use, or a message prompting them to try harder for daily use. Weekly, for the remainder of the first month, they received generic text ‘pop-ups’ on the application. For months 2-3, patients were then asked to do a short photo log of daily medication administration. For months 4-6, they were asked to maintain a log and continue to record videos if they desired. In addition, during the initial 3 months, patients participated in educational modules and quizzes related to SCD/thalassemia, iron overload and chelation therapy. Data was analyzed starting after one week of participation. Eight patients (4 males and 4 females, median age 12.5 years old, range 8-19) have been consented to participate. Seven patients have SCD and 1 has thalassemia major. We have analyzed 106 data days for 5 patients; Two patients have not yet reached 1 week of participation, and one patient has been temporarily excluded due to difficulties with internet connectivity and inability to confirm entries. Compliance of recording ‘selfies’ was 84%. Average length of ‘selfie’ videos was 12.7 seconds for male participants and 74.7 seconds for females (range 3 to 300 seconds). Videos reviewed also included: messages to providers, indications of difficulties with taste, and interaction with parents while preparing medication. Three patients have returned for 30-day follow up. Mean ferritin in these patients prior to enrollment was 1948.3 ng/ml (SD 758.4) and at one month follow-up was 1589.3 (SD 147.6). This study illustrates that mobile technology in the form of an interactive application is a feasible method that can be used to increase adherence to therapy plans, such as oral chelation therapy. We were able to accurately document adherence (84%) using our newly developed mobile app and are interested to see long-term compliance following completion of the study. Interestingly, we also report longer ‘selfie’ videos for females and may argue for gender specific recommendations for improving compliance to treatment plans. Detailed review of videos also revealed information that may further assist in compliance such as identification of improper mixing. Further expansion of this study and evaluation of laboratory markers (such as ferritin) at 3 months will additionally support the use of a mobile technology based intensive training program for patients with SCD. Efforts are also underway employing a similar strategy for hydroxyurea adherence. In summary, we believe mobile technology has become an integral part of health care management and has the potential to improve care for patients with chronic illnesses. Disclosures Shah: Novartis: Speakers Bureau.

BACKGROUND Sickle cell disease (SCD) is the most common hemoglobinopathy worldwide, characterized by chronic complications due to ongoing vaso-occlusion and hemolysis. Previous studies have shown that red cells from individuals with sickle cell disease (HbSS) have reduced NRF2 expression levels, which contribute to decreased oxidative stress capacity and increased hemolysis (Sangokoya, et al. 2010 Blood). Additionally, Macari and Lowry have shown that in vitro NRF2 activation of erythroid progenitors results in induction of anti-oxidant stress response genes, as well as increased percent fetal hemoglobin (HbF), which is known to prevent sickling (2011 Blood). Therefore, we hypothesize that NRF2 activation in SCD patients has potential therapeutic benefits by simultaneously inducing HbF and increasing the anti-oxidative stress capacity of red cells. We proposed to activate NRF2 by using sulforaphane (SFN), a well-known natural product enriched in broccoli sprouts. We conducted an open-label, dose-escalation clinical trial for SCD patients to investigate the safety and physiological effects of NRF2 activation by SFN through ingestion of a broccoli sprout homogenate (BSH). METHODS Male and female adult patients (> 18 years) with either HbSS or HbSߺ thalassemia were enrolled at the Duke Comprehensive Sickle Cell Center adult clinic. Exclusion criteria: RBC transfusion or a change in hydroxyurea dose in the last three months, ongoing pregnancy, diabetes, or renal insufficiency (BUN >21 mg/dL and/or creatinine >1.4 mg/dL). Inclusion criteria: Hematocrit (Hct) ≥ 20% and Hb > 6.0 g/dL. Recruited subjects were instructed to avoid additional SFN-containing foods before and during the study period. Subjects ingested a thawed preparation of BSH once daily for 21 days to allow for repopulation of red cells during therapy. Tolerability, toxicity, and physiological effects of NRF2 activation were determined at pre-treatment baseline (day 0), on the last day of ingestion (day 21), and after a wash-out period (day 49). Five patients were recruited for each dose (50g, 100g), with the smaller dose having elicited no safety concerns. RESULTS No safety concerns were noted among the subjects at either dose. In both cohorts, there were no significant differences in the adverse events, pain scores, complete blood counts, complete metabolic profile, reticulocyte count, and LDH levels when comparing days 0 and 21. In the 50g cohort, there was an overall but not statistically significant increase of average HbF from 14.5% to 14.9% (p=0.0786) in all five patients from Day 0 to Day 21; analyses are incomplete for the higher dose. We also observed a trend of NRF2 mRNA target gene induction, including heme oxygenase-1 (HO-1), NAD(P)H dehydrogenase (quinone 1, NQO1), and globin mRNAs, at day 21 vs day 0, which returned to baseline levels at day 49. In the 50g cohort, we observed a 66% increase of ho-1 and 44% increase of nqo1 mRNA levels at day 21 vs. baseline. In the 100g cohort, we observed a 14% increase of ho-1 and 42% increase of nqo1mRNA levels at day 21 vs. baseline. CONCLUSION Our pilot trial suggests that NRF2 activation by BSH may increase NRF2 expression programs and induce fetal hemoglobin. We aim to enroll more patients at escalating doses, as participants present a wide range of clinical variability and may show variable response. Additionally, the lack of statistical significance at the lowest doses along with a lack of safety concerns strongly compel us to pursue more potent NRF2 inducers to elicit more robust physiological changes for additional clinical trials. Disclosures No relevant conflicts of interest to declare.

Abstract 4698 Introduction: Information technology (IT) in general, and mobile technology in particular, is increasingly used in health care management. Mobile health tools have the potential to increase health care quality, reduce disease burden among individuals and health communities alike, as well as reduce healthcare disparities. Currently, there is a lack of data about ability and interest of patients with hemoglobinopathies, especially sickle cell disease (SCD) and thalassemia (Thal), to use IT interventions as part of their heath care management. This study was designed to ascertain the current use of IT, particularly mobile technology, in adult patients with hemoglobinopathies, as well as readiness to use mobile and other IT devices in the management of their disease. To do so, we sought to measure the ownership and use of IT devices (computers, mobile phones, and tablets) in a predominantly SCD population and the comfort level in using those devices to communicate socially and with health providers via text messages, emails, social media, or chat rooms. The study goal was to gather data pertinent to the potential use of mobile technology in clinical care, patient or provider education, and research interventions. Methods: The study consisted of a questionnaire-based telephone survey gathering patients' demographics and current use of IT, particularly mobile technology. Patients were also asked to rate their comfort level with computers and smartphones (0: not comfortable to 10: extremely comfortable) and their level of interest in the use of mobile devices for health care management. Using an IRB approved protocol, patients were randomly selected from the list of all patients that received care at the center in the preceding two years. Results: One hundred adult patients with hemoglobinopathy and longitudinal follow-up at the outpatient clinic were enrolled. All patients approached agreed to participate. More than 95% of patients had SCD; the rest had thalassemia or other hemoglobin variants. Fifty-seven percent were female. Forty percent were 18–34 years old, 34% were 35–50 years old, and 25% were older than 50 years. Ten percent (10%) of patients reported not having a high school diploma (HS-dip), 36% had a HS-dip without further education, 48% were college graduates, or had attended some college, and 6% were attending graduate school, or had achieved an advance degree. These four groups were regrouped into two (HS-dip or less vs. attended or graduated from college) for analysis. Eighty-four percent (84%) reported owning a computer device, i.e. desktops, laptops, tablets, iPads. Ninety-two percent (92%) reported having a mobile phone. Modes of communication were examined by age and education level. For all age and education groups, texting and emailing were preferred over social media, and particularly chat rooms (p Conclusion: This study suggests the feasibility of appropriate disease-specific health information technology (HIT) solutions for SCD clinical care, patient education, and patient-oriented research interventions Disclosures: No relevant conflicts of interest to declare.

Abstract 3230 Sickle cell disease (SCD) is caused by a single amino acid substitution of the β-globin chain of hemoglobin, resulting in abnormal red blood cells that occlude vessels, causing tissue damage, pain, and anemia. While all individuals with SCD have the same β-globin mutation, disease severity varies and is correlated with the concentration of retained fetal hemoglobin (HbF) (Akinsheye et al, 2011). After birth, a switch from γ- to β-globin gene expression occurs and HbF (α2γ2) is replaced by HbA (α2β2). In patients with SCD, increased levels of γ-globin can help compensate for the effects of the β-globin mutation. SCD patients with higher than normal retention of HbF frequently have more benign disease symptoms and longer life expectancy (Platt et al, 1994). Increased HbF expression through the use of hydroxyurea (HU) is a frequently used therapy and the only FDA-approved medication for SCD. However, due to the toxicity and inconsistent efficacy of HU, further understanding regarding the regulation of this “hemoglobin switch” is essential for developing alternative treatments. DNA methylation is central to the silencing of γ-genes during the switch to β-globin expression (Goren et al, 2006). Methylation marks are established by the DNA methyltransferase DNMT3A, which is recruited to the γ-promoter by the protein arginine methyltransferase, PRMT5 (Rank et al, 2010). We first hypothesized that genetic variation in DNMT3A and PRMT5 contributes to HbF levels. To test this, we performed a genetic association study on 603 unrelated, adult SCD patients. 21 haplotype-tagging SNPs in DNMT3A (n=17) and PRMT5 (n=4) were genotyped in the patients. SNPs were evaluated for association with HbF levels, as well as the occurrence of severe pain and narcotic use, which were used as surrogates for non-hemoglobin effects of HU. HbF levels were measured when the patients were at steady state and not taking HU. Severe pain was defined as a history of hospitalization for pain within a 12 month period. Narcotic use was defined as taking long- or short-acting narcotics to alleviate pain at home. To evaluate associations between the SNPs and clinical outcomes, we used linear and logistic regression (SAS Systems, Cary, NC), while controlling for age, sex, degree of European ancestry and HU treatment. European ancestry was estimated from 4331 genome-wide ancestry informative markers (Ashley-Koch et al, 2011). HbF levels were log-transformed prior to analysis in order to achieve a normal distribution. We corrected for multiple comparisons using the Li and Ji method (Li & Ji, 2005). Two SNPs in DNMT3A were significantly associated with the occurrence of severe pain and HbF levels after correcting for multiple testing. Additionally, two SNPs in DNMT3A and one in PRMT5 were nominally associated with severe pain, two SNPs in DNMT3A were nominally associated with narcotic use, and one SNP in PRMT5 was nominally associated with HbF levels. These findings demonstrate that genetic variation in key genes responsible for the regulation of DNA methylation at the γ-globin locus is associated with SCD clinical outcomes. We next hypothesized that the variability in clinical outcomes might be due to epigenetic variation at the γ-globin locus. In order to determine specific DNA methylation patterning of γ-globin genes (HBG1 and HBG2), we performed pyroseqencing subsequent to bisulfite treatment of DNA from a subset of 72 SCD patients, all of whom were HbSS, and 50% of whom were not on HU treatment. Percent methylation at the γ-globin locus was nominally associated with HbF levels among patients not taking HU (p=0.03), but not among patients taking HU. Additionally, we investigated whether SNPs predicted methylation status and identified two SNPs in DNMT3A (rs734693 and rs7583409) to be predictive of methylation status at the γ-globin locus. These data provide further insight into the complex regulation of the γ-globin locus and suggest that genetic variation in DNMT3A and PRMT5 is associated with clinical outcomes and methylation status in SCD patients. Future studies are needed to further investigate the impact of epigenetic processes as a potential mechanism for HbF expression and induction. Disclosures: Telen: GlycoMimetics: Research Funding.

Abstract 1648 Background: Renal failure occurs in 5–18% of sickle cell disease (SCD) patients and is a major risk factor for early mortality. However, there is no established method of identifying SCD patients that are at high risk of developing this outcome prior to the appearance of proteinuria, and its pathobiology is not well understood. The non-muscle myosin heavy chain ll-A (MYH9) gene, which encodes the heavy chain of myosin II-A in the podocyte cytoskeleton, has been identified as driving the high risk of focal segmental glomerulosclerosis (FSGS) and end-stage renal disease in African Americans. Methods: We genotyped 26 single nucleotide polymorphisms (SNPs) in the MYH9 gene in 521 unrelated adult (18 – 83 years) SCD patients who had been screened for proteinuria. Logistic regression was used to determine if the SNPs predicted risk for proteinuria among the patients. Results: Of 521 adult SCD patients studied, 140 had proteinuria, while 381 did not. On average, subjects with proteinuria were 6 years older than subjects without proteinuria (p Conclusion: Our data provide additional support for the role of MYH9 in renal dysfunction among African Americans. A specific haplotype appears to be associated with increased risk for proteinuria among patients with SCD. The association of MYH9 with renal dysfunction in SCD provides insight into the pathophysiology of this process and may lead to early identification of patients at risk and, ultimately, to new modes of therapeutic intervention. Disclosures: De Castro: GlycoMimetics: . Telen:GlycoMimetics: Consultancy, clinical trial sponsorship.

Abstract 1529 Poster Board I-552 Pulmonary hypertension (PAH) is an independent risk factor for death in sickle cell disease (SCD). We performed a retrospective chart review to determine the natural history of PAH in our adult SCD population. We hypothesized that increased pulmonary artery pressures seen during hospitalizations for painful crisis or acute chest syndrome would be reflected in a faster progression of PAH measured during steady state. We reviewed charts and echocardiograms of 362 patients seen at Duke University from January 1980 to March 2009. A total of 878 2D echocardiograms were reviewed, with 196 patients having 2 or more echocardiographic procedures. Fifth-three of the 81 patients who died during this period had had at least 2 or more echocardiograms. Studies were considered done at steady state if they were performed as an outpatient procedure. Out of 878 total echocardiograms, 460 had either no measurable tricuspid regurgitation jet (TR) or failed to report it and were excluded from further analysis, leaving 418 echocardiograms in 252 patients (167 at steady state and 251 as inpatient). Among patients with multiple echocardiograms and measurable TR jet performed at steady state, we used a linear mixed model to identify a mean yearly rate of progression of 0.04 m/s (p Disclosures No relevant conflicts of interest to declare.

Pulmonary artery hypertension (PAH) occurs in 30–50% of adult patients with sickle cell disease (SCD), with mortality ranging from 16 to 50% and a median survival of 25 months. Our objective was to use gene expression profiling to develop a gene signature predictor for PAH through the analysis of gene expression of blood cells from SCD patients with or without PAH. We hypothesized that these gene signatures could allow us to identify patients at risk for PAH, as well as to generate hypotheses as to the pathophysiology of PAH in SCD. We used Affymetrix U133A2 GeneChip to determine the RNA expression of both whole blood and leukocytes using PAXgene and Leukolock methods, respectively. The study population included patients homozygous for HbS or with HbSβ0 thalassemia. Subjects with PAH were ≥18 years old, in steady state, and had PAH either by 2D echo (TR jet ≥ 2.7 m/sec) or right-sided catheterization (mean PA pressure ≥ 30 mmHg). Patients were excluded if they were pregnant, had co-existing rheumatologic conditions or other inflammatory diseases, were on chronic transfusion therapy or had had a vaso-occlusive episode in the previous 4 weeks. The control subjects were patients with SCD but without PAH (TR jet ≤ 1.8 m/sec or mean PA pressure Figure Figure

Many of the tertiary care hospitals in North Carolina (NC) are often frequented by patients who have to travel a long distance, given the rural nature of the state. Nearly one half of the adult sickle cell disease (SCD) patients seen at the Duke Comprehensive Sickle Cell Center (DCSCC) come from areas further than a 1-hr drive. The current study aims to determine whether geographical proximity to a comprehensive medical center is associated with SCD outcomes, as indicated by severity score, hospitalization frequency, and quality of life. Methods: Two hundred and two patients who primarily receive their SCD disease-related care from DCSCC were enrolled in the study. The sample included 101 males and 101 females, aged 20–69 years (mean=35.6), with SCD disease (SS: n=135; SC: n=47; other: n=20), and level of education ranging from 4–18 years (mean=13.1). Patients lived an average of 50.4 miles (median: 38.6, range:0.4 to 383) from DCSCC. Linear regressions, controlling for age and SCD diagnosis, were used to test associations between continuous variables. Severity scores measuring end organ damage were determined as previously described (Afenyi-Annan et al. 2008), and frequency of hospitalizations over the past 2 years was determined by self-report and medical record review. To measure mental and physical quality of life (QoL) domains, patients were administered the SF36 QoL scale. Hydroxyurea (HU) and opiate pain therapies were also recorded. Patients were considered to be on opiates if they had used opiates daily for a period of thirty days in the previous 12 months. Results and Discussion: Living closer to Duke had a statistically significant association with higher disease severity scores (β = −0.17, p=0.01). Moreover, proximity to Duke was associated with higher frequency of hospitalizations (β = −0.23, p=0.002). These associations were not modified by gender, employment status or education. Medication use did not account for the association between proximity and disease severity, or proximity and frequency of hospitalizations. The mental domain scores of self-reported QoL correlated negatively with hospitalizations (r= −0.18, p=0.02), whereas the physical domain score negatively correlated with both disease severity (r= −0.19, p Conclusion: Patients who live closer to our tertiary care comprehensive center had higher disease severity scores and more hospitalizations over a two year period than patients who live farther away. Neither age, disease diagnosis, gender, employment status, education, nor HU and/or opiate medication use accounted for the negative association between proximity to DCSCC and disease outcomes. On the other hand, distance from DCSCC did not affect patients’ quality of life. The cross-sectional nature of the current study makes it difficult to determine causality. However, it is possible that patients who live close to a major medical center rely more on health system availability as a means to managing their disease, while those living further away rely on self-care at home or adhere to long-term medical regimens. Health care providers may need to focus on developing practice guidelines that encourage and empower patients to take a more active role in their medical care and be less dependent on their healthcare providers to decrease frequency of hospitalization and, perhaps decrease the progression of their disease.

Pregnancy in sickle cell disease (SCD) has been associated with complications and adverse outcomes for mother and child and thus warrants specialized clinical care. There is bidirectional impact between pregnancy and SCD. Only a small number of previous studies of pregnancy outcomes in SCD patients have been published, and some have been based on nationwide diagnosis queries rather than direct patient queries. An increased incidence of spontaneous abortion, pregnancy-induced hypertension, infections, pre-term labor, and low birth weight was noted in those reports. The aim of the present study was to identify and describe the characteristics and outcomes of pregnancy and other gynecological events in our current patients with SCD. We hypothesized that these women experience a higher frequency of pregnancy-related complications and earlier onset of menopause than the general population. One hundred adult female SCD patients from the Duke University Medical Center sickle cell clinic were included in this study. Sixty five were homozygous for hemoglobin (Hb) SS, 22 had Hb SC and 13 had other Hb genotypes. A standard questionnaire was developed, and patients were interviewed either personally or by telephone after obtaining IRB-approved informed consent. Most of the data reported here comes from patient interviews. Medical records were reviewed only to confirm Hb genotype and age. The mean age of individuals included in the study was 38.7± 13 years (range 19–75 yr). Sixty-seven of 100 reported a history of regular menses. The median age at menopause was 45 yr (range 27 to 58) as compared to 49.3 yr in the general African American population (AA) (Bromberger, Am J Epidemiol. 1997). Twenty-five percent of the patients were never pregnant, 92% of those (23 out of 25) stated they did so by choice. Seventy-five patients reported a total of 158 pregnancies; there were 111 live births and 3 still-births. One tubal pregnancy was terminated. Patients reported a total of 43 abortions: 28 were spontaneous, and 15 were induced. Twenty-seven (36%) patients have had at least one abortion. Of women with a history of pregnancy, 57 (76%) reported unplanned pregnancies. Mean gestational age was 35.9 ± 5 weeks. Maternal complications evaluated included: thrombosis in 6 patients (8%), five in the lower limb and one in the lungs; and pregnancy-induced hypertension in 10 (13%) patients. The rate of live births was 0.7. The prevalence of low birth weight was higher than in the AA population (26.3% vs. 13.2%, respectively) and the mean birth weight was lower than the AA population (2595.5 vs. 3089 g, respectively) (David et al, NEJM. 1997). The mean birth weight for women who increased their use of pain medications during pregnancy was lower than that for women who continued on the same pain medication regimen, those who took less medication, and those who did not use pain medication (2045, 2743, 2758, and 2924 g, respectively, p < 0.02). The babies of patients in the first group were more likely to stay longer in the hospital (p = 0.006). We conclude that obstetrical and gynecological outcomes in SCD patients differ significantly from and are in general worse than for the AA population, especially for menstrual history, onset of menopause, prevalence of low birth weight and mean birth weight. Moreover, increased use of pain medications in pregnancy was associated with lower birth weight. This finding is troublesome and deserves further study.

The six-minute walk test (6MWT) is a standardized test use to measure patient cardiopulmonary functional capacity. It is been increasingly used as a measure of overall “functional ability” in chronically ill subjects. The frequent occurrence in sickle cell disease (SCD) of complications such as severe anemia, hypoxemia, and avascular necrosis (AVN) raises questions regarding the capacity of this patient population to perform the 6MWT adequately. This study prospectively evaluated the performance of SCD patients enrolled in a longitudinal prospective study evaluating pulmonary hypertension (pHTN) natural history in a 6MW test. The 6MWT was performed in adult patients with SCD, at steady state. The following 6MWT variables were analyzed: Hemoglobin (Hb) diagnosis, age, Hb level, gender, heart rate (HR) at rest and maximum predicted rate at exercise, distance walked (DW), and percentage (%) of predicted distance walked (%PDW), pulse oxymetry (O2Sat) at rest and at exercise, Wellness scale, Borg scale, reason for resting, and physical limitations in performing the test. In addition, pHTN was evaluated by 2D echo. Patients with a TR jet ≥2.5 m/s were considered to have pHTN. Data from 52 adults (≥ 18 yr, 21 females) were collected and analyzed. Thirty-two (62%) had Hb SS, 13 Hb SC, 6 Hb Sβ+Thal, and 1 Hb SδβThal. TR jet was measurable in 41 patients, and 16 of them (39%) had a TR jet ≥2.5 m/s (range 2.5 – 4.7). Mean age of subjects was 36.5 ± 12.6. Females were older than males. (41.3 ± 2.8 vs. 33.7 ± 2.0; p=0.03). The mean distance walked by all patients was 463.6 meters ± 99.2 (minimum 142, maximum 658). The mean %PDW was 72.07% ± 20.75, range: 24 to 146%. There was no statistical difference in the DW when the different HbS genotypes were compared. As expected, the mean DW was higher in males than females (488 ± 14.4 vs. 427.4 ± 25.1 meters, p

Hydroxyurea (HU) is used to treat sickle cell disease and has been shown to decrease painful episodes (Charache, 1995) and possibly vaso-occlusive episodes associated morbidity and mortality (Steinberg, 2003). Opioids are often prescribed in adult patients for daily management of their chronic pain. The aim of the current study was to determine the age at death and the effect of treatment with HU and/or opioids prior to death in patients who died from sickle cell disease (SCD) related complications and to compare these parameters to those in our current patients population. Methods: Age, daily treatment with opioids, and HU treatment were determined for 185 patients currently followed at the Duke Comprehensive Sickle Cell Center (DCSCC) and for 50 patients who died between 2002 and 2008 due to SCD complications and who were regularly followed at the DCSCC for their care. The two cohorts, living and deceased patients were divided based on their treatment modality into the following 4 groups: opioid only, HU only, both drugs, and neither drug. Non-parametric chi-square test was performed to determine whether the treatment modality distribution was different in the deceased group compared to the living group. Analysis of variance was done to determine the relationship between treatment group and age at death. Results and Discussion: The distribution of treatment modalities in the deceased group was significantly different than that of the living group. The opioids only group had the largest number of patients in the deceased cohort (44%), and this percentage was almost twice that of the living group (25%). (Fig. 1) Moreover, 72 % of the deceased patients were treated with opioids vs. only 53 % of the living patients, perhaps because the sicker patients are often treated with daily opioids. However, the age of death in the opioids only group was 44 ± 15.5 years. (Fig. 2) In the living group, treatment with both drugs or with no drugs were equivalent (28%), while in the deceased group, more patients were treated with both drugs (28%) compared to no drug treatment (18%). The HU only group had the lowest number of deaths (10%), and the percentage of patients in this group was nearly half that the one of the living group (19%). This group also was the oldest at death (58 ±16 years). Age at death was also significantly higher in this group than in each of the other 3 groups (p

In 1994, the average age of death of patients with sickle cell disease (SCD) in the US was 42 years for males and 48 for females. We theorized that patients who lived appreciably beyond these ages would therefore have lower disease burden than patients overall, and that lack of specific disease complications would be characteristic of such patients. From 2001 until 2006, we enrolled 541 unrelated adult patients (age ≥18) with Hb SS or Hb Sβ0 at three comprehensive sickle cell centers in a study designed to identify factors associated with clinical outcomes in SCD. Our database includes demographic, clinical, and laboratory data on all participants. We identified 61 patients 50 years and older and 32 patients 55 years and older. Among the latter, the average age was 60.6, with a median of 58 years (range 55–83); 65% were female. Although patients with SCD are generally thought to have a low prevalence of hypertension, the mean BP in this older cohort was 142/75, and 55% of patients were taking anti-hypertensives. Patients ≥55 had the following history of SCD-related complications: acute chest syndrome – 69%, priapism (males only) – 46%, stroke – 16%, TIA – 10%, seizure – 3%, AVN of shoulders or hips – 40%, leg ulcers – 38%, heart failure – 16%, gallstones – 64%, and retinopathy – 40%. Compared to all patients, elderly SCD patients had higher prevalence of TIA (10% vs 5%), retinopathy (40% vs 21%), and heart failure (16% vs 6%), and lower prevalence of seizures (3% vs 12%). When compared to younger patients, the elderly had a similar rate of AVN (40% vs 30%) but a significantly higher number had undergone surgical joint interventions (33% vs 10%; p=0.0001). The percentage of patients who had had cholecystectomies and splenectomies was uniform through the whole study cohort. Most interestingly, 60% of patients ≥ 55 had findings consistent with pulmonary hypertension, defined as TR jet ≥ 2.5 m/s on echocardiography. This represented approximately twice the prevalence seen in the entire study cohort. Significant proteinuria (≥ 1+) was present in 41% of patients, again approximately twice as frequently as seen in younger patients. Interviews and review of records showed that 47% of elderly patients had not required any hospitalizations during the past year (compared to only 27% in the total study cohort), while 20% were hospitalized only once, 20% were hospitalized 2–4 times, and 6% were hospitalized >4 times. Review of medication usage revealed that 35% were taking hydroxyurea (HU) at the time of enrollment, and 13% used long-acting narcotics daily (compared to 39% and 24% of the total cohort, respectively). When hematologic parameters were compared for elderly patients who were or were not taking HU, no significant differences were observed for Hb, Hct, WBC and platelet counts, although the mean Hb for patients taking or not taking HU were 8.5 and 7.8 g/dL respectively. We conclude that when compared to the overall population, the elderly had a higher prevalence of pulmonary hypertension, systemic hypertension, TIA and joint replacement, suggesting that contrary to our hypothesis, the degree to which they had experienced SCD-related end-organ damage was not decreased. While the frequency of HU use was similar to all patients, HU did not have clear effects on hematologic parameters in the elderly, raising the question of its efficacy in this population. Finally, the elderly did have some indicators of lower disease severity—namely, lower use of daily long-acting narcotics and fewer admissions for painful episodes. While research has traditionally focused on the high-risk SCD population, we believe that further investigation of elderly patients as a “lower risk” group is warranted to further advance our understanding of the relationship of SCD complications to survival.

Sickle cell disease (SCD) patients experience high rates of morbidity and early mortality. Hydroxyurea (HU) therapy is associated with decreased morbidity and mortality as well as improved patient health outcomes (Charache et al, 1995 and Steinberg et al, 2003). However, the effect of chronic opiate pain management on HU treatment outcomes is unknown. In the current study, we evaluated the effects of HU and chronic long acting opiate therapy-OxyContin, methadone, fentanyl and MS Contin-on the following laboratory parameters: white blood cell count (WBC), hemoglobin (Hb), and mean corpuscular volume (MCV). Changes in these variables are known to be linked with HU effects on morbidity and mortality in SCD. 142 homozygous sickle cell anemia patients (75 females and 67 males) from the outpatient clinic at Duke University Medical Center were included in this study. Clinical data were collected by patient interview as well as review of medical records after informed consent. Patients were classified in 4 groups based on report of chronic daily use of long acting opiates and/or HU (current therapy with HU) as follows: No HU or opiates (None); opiates only; HU only; and HU and opiates. ANOVAs were used to identify combined effects of opiate therapy, HU therapy and the 3 laboratory parameters. Overall, we found that patients on HU (n=76, 54%), had lower WBC and higher Hb and MCV than the no HU patients (n=66, 46%), with p Figure Figure

Genetic risk factors may contribute to the incidence of certain sickle cell disease (SCD) complications. Priapism, a painful, unwanted and sustained erection, is caused by decreased venous drainage in the penile vessels. Our previous work suggested an association between multiple non-coding polymorphisms in the A1 subunit of coagulation Factor XIII (F13A1) gene and a history of priapism (Elliott et al., 2007). The purpose of this study was to further investigate the possible association of polymorphisms in the A1 and B (F13B) subunits of the Factor XIII gene with priapism in order to better understand how differences in the coagulation pathway might affect the risk for priapism. The larger data set from which this study was derived included 524 unrelated patients with any form of SCD. These individuals had been recruited at three southeastern US medical centers. A subset of 186 males from this study population was used in the current analysis. These patients all had a diagnosis of Hb SS, had provided a DNA sample for genetic analysis and had responded to a standardized questionnaire regarding the history of priapism. The patients had a mean age of 32.4 years, and 78 (42%) reported a positive history of priapism. Four coding single nucleotides polymorphism (SNPs) in the F13A1 gene and 10 SNPs (2 coding, 8 non-coding) in the F13B gene were genotyped in our dataset. Genotyping was performed using Applied Biosystems Taqman Allelic Discrimination Assays. For quality control, blinded duplicate and CEPH samples were included on all gels and required to match 100%. Further, the genotypes of at least 95% of the samples had to be called with certainty to be considered for statistical analysis. Deviations from Hardy-Weinberg equilibrium (HWE) were identified using exact tests implemented by the Genetic Data Analysis program; no markers in F13A1 or F13B significantly deviated from HWE. For each SNP, exact tests of association for the genotypes by occurrence of priapism were performed using SAS. No SNPs in F13B were associated with priapism, and two F13A polymorphisms previously associated with risk for thrombosis (rs5985 and rs6003) were also not associated with risk for priapism in our data set. However, a single coding SNP in F13A1 (rs5988) was strongly associated with priapism (global p-value of 0.03). The odds that males with the C/C genotype suffered from priapism were 2.43 times higher than for those with the C/G genotype. This SNP resides in the C- terminus of FXIIIA. This area of the protein functions to prevent substrates from gaining access to the Factor XIII active site. While there is no evidence that this area directly regulates catalysis, it may play a role in aligning the substrate with the active site. Factor XIII could modify fibrin formation and alter red cell interaction with the vasculature. We plan to express the molecule encoded by the allele associated with risk to see if it has altered activity. In conclusion, identification of risk groups for priapism by genetic studies may lead to investigation of newly identified pathophysiologic mechanisms for this complication of SCD. | | Negative History | Positive History | Total | |:----- | ---------------- | ---------------- | ----- | | C/C | 68 | 61 | 129 | | C/G | 38 | 14 | 52 | | G/G | 2 | 3 | 5 | | Total | 108 | 78 | 186 | Priapism Genotype Frequencies for rs5988

Pain is a limiting factor in the daily life activities of sickle cell disease (SCD) patients. Opioid analgesics are widely used for management of SCD-related chronic pain in the community. To date there have been no studies on the use of opioids and their impact on quality of life (QoL) measures in this population. Patients on long-term opioid pain management may also be on hydroxyurea (HU), which is often used to decrease the frequency of painful episodes and other SCD sequelae. HU is also known to affect some QoL measures. The aim of this study was to determine the effect of opioid analgesia on QoL measures in an adult SCD population. This study included 185 patients, 92 females and 93 males, from the outpatient clinic at Duke University Medical Center. 117 patients either had homozygous SCD or were doubly heterozygous for Sβ0 thalassemia; 68 patients were of different genotypes. Data were collected by patient interviews as well as review of medical records after informed consent. The Medical Outcome Study 36-item Short Form (SF-36) was used to determine QoL measures, and the results were scored in standard fashion. Differences in variables of interest between narcotic users and narcotic non-users were analyzed using t-tests. ANOVAs were used to identify combined effects of narcotics, HU and their interaction with SF-36 scores. Gender, age and genotypes were included as covariates. Patient data were classified in 4 groups based on report of regular use of opioids (at least 30 days within the preceding year) and HU (any use during the preceding year) as follows: no HU or narcotics (none); narcotics only; HU only; and narcotics and HU. SF-36 scores for all physical and mental domains were significantly lower in individuals on opiates vs those not on opiates, in all age groups (p Figure Figure

Sickle red blood cells (SS RBC) are abnormally adhesive to both endothelial cells (ECs) and components of the extracellular matrix (ECM). Epinephrine (epi) has been shown to elevate cAMP in SS RBC and increase adhesion of SS RBC to ECs in a protein kinase A-dependent manner. In vitro and in vivo studies performed in our lab have led to the hypothesis that adrenergic stimuli such as epi may initiate or exacerbate vaso-occlusion and thus contribute to the association of vaso-occlusive events with physiologic stress. We are conducting a prospective, dose-escalation pilot clinical study to investigate whether in vivo administration of one dose of propranolol either down-regulates baseline SS RBC adhesion in vitro or prevents its upregulation by epi. In addition, this study will provide additional safety data regarding the use of propranolol in normotensive patients with sickle cell disease (SCD). Figure Figure To date, we have completed the first two dose cohorts. 11 subjects (9 SS and 1 Sβ° thalassemia; 7 females, 3 males) have participated. No severe adverse events were noted. Cohorts 1 and 2 had mean pre-propranolol blood pressure (BP) of 116 (5.9 SD)/ 60.4 (3.98 SD) and 106.8 (4.68 SD)/ 58 (3.9 SD), respectively; this difference was not statistically significant. Minimal and asymptomatic changes in BP were noted in both cohorts after drug administration, with biphasic systolic and diastolic BP nadirs at 45 and 240 minutes. No clinically significant changes in heart rate were observed. Adhesion studies were performed using a graduated height flow chamber on the day of RBC collection. RBC adhesion to ECs was studied before and after epi stimulation and was measured at sheer stresses ranging from 1 to 3 dyne/cm2. Baseline adhesion measurements were validated by comparing percent (%) adhesion assayed at 2 different times within 7 days—at screening and before propranolol dose on the study drug day. We observed no significant difference in adhesion at the 2 different time points without propranolol. Comparison of % adhesion of epi-stimulated RBC to ECs before and 1 hour after propranolol showed that propranolol given in vivo significantly inhibited both non-stimulated and epi-stimulated SS RBC adhesion (p=0.04 and p=0.001, respectively). Lastly, comparison of SS RBC adhesion at both drug doses confirmed the drug-related inhibition of adhesion (p

Over the last five years, we have enrolled 541 unrelated adult patients (age >18) with HbSS or HbSβ0 at three major comprehensive sickle cell centers in a study designed to identify factors associated with clinical outcomes in sickle cell disease. Our database includes demographic, clinical, and laboratory data on all participants. Medical history was obtained by both patient interview and review of medical records. The average age of patients enrolled was 34.0 yrs, median age 31.9, and 61 were ≥ 50 yrs; 54% were female. 81% of patients had finished high school, and 35% smoked tobacco products. History of SCD-related complications was as follows: acute chest syndrome - 76%, priapism (males only) - 41%, stroke - 16%, TIA - 5%, seizure - 12%, AVN of shoulders or hips - 30%, leg ulcers - 26%, heart failure - 7%, gallstones - 66%, and retinopathy - 22%. 68% of patients had undergone cholecystectomy, 11% splenectomy, and 11% major joint replacement. 136 patients had echocardiography data reported, and of these 39% had findings consistent with pulmonary hypertension, as defined as TRjet ≥ 2.5 m/s. 7% were receiving chronic transfusion therapy, and 6% were receiving iron chelation therapy. Significant proteinuria (≥ 1+) was present in 27% of patients, while only 4% had hematuria. Review of medication usage revealed that 40% were taking hydroxyurea (HU) at the time of enrollment, and 23% used long-acting narcotics daily. 14% of patients took medication for hypertension, 9% took antidepressants, and 6% used bronchodilators. Interviews and review of records showed that 21% had not required hospitalization during the past year, while 25% were hospitalized only once, 46% were hospitalized 2–4 times, and 8% were hospitalized >4 times during the past year. Hematologic values and HU status were available for 433 patients. Hematologic Values On HU Not On HU Mean Std Dev N Mean Std Dev N p value Hb 8.9 1.62 174 8.29 1.46 259 Patients not on HU had significantly higher total bilirubin values (3.51±2.66 vs 2.91±2.93, p=0.02) and insignificantly higher ferritin levels (1297±2118 vs 1089±1535, p=0.5). However, patients taking HU had higher LDH (p=0.0019) and uric acid levels (p

Sickle cell disease (SCD) is associated with extensive morbidity and early mortality. Although the most common known causes of death for adults with SCD are acute chest syndrome, stroke, pulmonary hypertension, and infection, the direct cause of death is frequently undefined, and patients often die suddenly. In one series of 306 autopsies of patients with SCD, death was sudden and unexpected in 41% of cases (Manci et al 2003). The incidence of sudden cardiac death and associated risk factors in patients with SCD are currently unknown. We sought to identify risk factors for mortality in adult subjects with SCD and to evaluate the frequency, risk factors and co-morbidities of sudden death in this population. We identified 43 adult patients (21 males and 22 females) who had been followed in the SCD clinic at Duke University Medical Center (DUMC) and who had died between January 2000 and April 2005. Clinical characteristics and laboratory data were evaluated by retrospective chart review. Findings were compared with data from patients who were actively followed during the same time period and were still living (n=197). The average age at death was 44.3 years (range 21–83). The most frequently listed causes of death were liver failure, multiorgan failure, stroke, and pulseless electrical activity (PEA) arrest. The etiology of death in 29 of the 43 patients was unknown. Recognized risk factors for sudden cardiac death, including ejection fraction (52% vs. 54%), left ventricular size (LVIDd 5.0cm vs. 5.2cm), and fractional shortening (0.30 ±0.01 vs. 0.33± 0.01) as measured by echocardiogram, were not significantly different between deceased and living patients, respectively. Left ventricular hypertrophy (LVH), defined as a left ventricular mass index ≥134 and ≥110 g/m2 for men and women, was reported in 41% of the deceased patients but in only 31% of living subjects. Of the 12 deceased patients with LVH, 7 had mild LVH and 5 had moderate-severe LVH. The average tricuspid regurgitant jet velocity measured by Doppler echocardiogram was higher in patients who died compared to those who were still living (3.72 vs. 2.17 m/s). The most frequently documented cardiopulmonary complications among deceased patients were acute chest syndrome/pneumonia, pulmonary hypertension, systemic hypertension, and stroke. Identified risk factors associated with premature death were pulmonary hypertension (p

The moderate to severe anemia associated with sickle cell disease (SCD) causes a high output state that can lead to cardiomyopathy, decompensation and left heart failure. While at least one report suggested that chronic high output and volume overload are relatively well tolerated in SCD, other studies have suggested that congestive heart failure and other cardiac abnormalities may occur in more than 50% of adults. The incidence of sudden death is also increased and may be as high as 40% in adults with SCD. Several types of left ventricular (LV) dysfunction are associated with sudden death in other populations, although an association of LV disease and sudden death in SCD has not been studied. For these reasons, and also based on the clinical observation that SCD patients frequently experience transient episodes of clinical congestive heart failure (especially when receiving intravenous fluids during vaso-occlusive episodes), we sought to document the frequency with which left-sided cardiac dysfunction existed in our patient population. We performed a retrospective review of resting echocardiograms to quantitate multiple parameters of cardiac function in patients either homozygous for hemoglobin S or with Sβ thalassemia. 116 patients had had at least one echocardiogram performed (55 females, mean age 37, range 13–71; 61 males, mean age 32, range 16–62). 31% of these patients (mean age 41) had left ventricular hypertrophy (LVH), defined as a left ventricular mass index ≥134 and ≥110 g/m2 for men and women, respectively, and 22% (mean age 35) had left ventricular enlargement. Of the 36 patients with LVH, 27 had mild LVH, and 9 had moderate-severe LVH. Presence and severity of LVH were significantly related to mean arterial blood pressure (p=0.004), even though most patients were not hypertensive. Mean arterial pressures were also related to posterior LV wall thickness (p

Sickle red cell (SS RBC) adhesion is believed to be a major factor in vaso-occlusion. SS RBCs adhere especially avidly to the extracellular matrix protein laminin (LAM), which is found in the subendothelial basement membrane as well as soluble in plasma. We asked (1) Whether the level of soluble LAM in plasma is related to chronic complications of SCD; and (2) Whether soluble LAM enhances the ability of SS RBCs to adhere to endothelial cells (ECs). We first established an ELISA assay to quantitate plasma LAM and found that plasma from all normal subjects tested had LAM levels ≤0.5 μg/mL, which was comparable to the lower limit of detection for our assay. However, plasma from HbSS patients in steady state had levels ranging from ≤0.5 to >8.1 μg/mL. Of 52 HbSS patients studied while in steady state, 5 had plasma LAM levels ≤0.5 μg/mL, 22 had levels >0.5 and 3.0 μg/mL) did not have a higher frequency of stroke, acute chest syndrome, priapism, leg ulcers, or aseptic necrosis than patients with levels 8.5 g/dL had LAM levels of 1.72±0.41 μg/mL (n=18, p=0.008). To evaluate whether soluble LAM contributed to SS RBC adhesion, we assayed adhesion using TNF-activated human umbilical vein endothelial cells in a graduated height flow chamber. When SS RBCs were first preincubated with physiologic amounts of LAM, we saw a marked increase in SS RBC adhesion to ECs, and this effect was related to the amount of LAM added. With addition of 5 μg/mL LAM, adhesion to ECs at 2 and 5 dynes/cm2 was increased by 20% and 46% over baseline, respectively. Addition of 10 μg/mL LAM enhanced adhesion at these shear stresses by a mean of 154% and 249%, respectively (n=4). Furthermore, SS RBCs from patients with lower plasma LAM levels had lower baseline adhesion in the absence of added LAM. We then incubated SS RBCs with ABO-compatible SCD plasmas with either high or low LAM levels, washed these cells, and tested them for their ability to adhere to ECs. Plasmas with high LAM (n=3, LAM 4.5–8.1 μg/mL) strongly enhanced adhesion by a mean of 71.9% over baseline at 2 dynes/cm2, while those with low LAM levels (0.4–0.6 μg/mL) enhanced adhesion by only 21.1%. Finally, in order to show that it was the LAM in these plasmas that enhanced SS RBC adhesion, rather than another factor elevated in parallel with LAM, we precleared high LAM plasma with monoclonal antibody reactive with LAMs-10,-11 and anti-murine Ig-Sepharose beads and then compared the activity of this plasma to the same plasma sham precleared with only Sepharose beads. Plasma LAM levels were also assayed to confirm removal of LAM by this procedure. Plasmas from which LAMs-10,-11 had been removed failed to enhance SS RBC adhesion, while the same plasmas subjected to sham preclears increased SS RBC adhesion to ECs by 76.9% at 2 dyne/cm2 (n=2). Thus, we conclude that plasma LAM can enhance SS RBC adhesion to ECs, and elevated levels of plasma LAM may contribute to vaso-occlusion. In addition, the inverse correlation of plasma LAM levels to Hb levels further suggests a relationship between SS RBC adhesion and the severity of anemia in SCD.

Priapism is a complication of sickle cell disease (SCD) that occurs due to obstruction of the corpora cavernosa of the penis. We have studied priapism in relation to several clinical and genetic factors in 249 adult male patients with SCD, 92 (37%) of whom reported a positive history of priapism. The mean age of male patients without a history of priapism was 35.2 years (± 10.8 years) compared with a mean age of 36.4 years (± 11.3 years) in male patients with a positive history of priapism. Because of the possible relationship with nitric oxide biology, we examined the co-occurrence of priapism with proteinuria, leg ulcers and stroke. Of the males with a positive history of priapism, 20% also had a history of 2+ or greater proteinuria, compared to a presence of 2+ or greater proteinuria in only 10% of males without a history of priapism (p=0.03). Similarly, 34% of males with a positive history of priapism also had a history of leg ulcers, compared to the presence of leg ulcers in 22% of males without priapism (p=0.03). No statistically significant association between the occurrence of priapism and stroke was observed. In an effort to identify genetic risk factors for priapism, we examined 262 single nucleotide polymorphisms (SNPs) in a total of 56 genes, primarily involved in red blood cell adhesion and inflammation pathways. Chi Square tests of association were constructed for the genotypes of each SNP with two clinical categories: patients with a positive history of priapism and patients without a history of priapism. When the frequency of rare homozygotes was less than five individuals, we combined these rare homozygote individuals with heterozygote individuals for analysis. All p-values were uncorrected for multiple testing. We found associations with 12 SNPs in 8 genes: SLC4A2 (p=0.003); ITGAV (p=0.004 and p=0.02 for two different SNPs); F13A1 (p=0.004 and p=0.02 for two different SNPs); AQP1 (p=0.01 and p=0.04 for two different SNPs); TGFBR2 (p=0.01 and p=0.02 for two different SNPs); ADRB2 (p=0.03); MGC (p=0.04); and ARG2 (p

The life expectancy of many patients with sickle cell disease (SCD) is well into the 5th and 6th decade, but this remains extremely variable. Little is known about the biological factors that protect certain SCD patients from early demise while others never reach mid-adulthood. Recently, McKerrell and colleagues (2004) compared the clinical and laboratory profiles of SCD patients aged 40 years and over with SCD patients who were between 21 and 30 years of age. Similarly, we have compared clinical and genetic correlates of older SCD patients (50 years and over) with those of younger patients (18–30 years). Among 514 patients in our total study population, 49 (10%) were categorized as “older” and 194 (38%) were categorized as “younger.” Older SCD patients had lower hemoglobin (older: 7.8 ± 1.1 vs. younger: 8.5 ± 1.2, p=0.004), platelet count (older: 372 ± 126 vs. younger: 460 ± 225, p=0.02), MCV (older: 92 ± 12 vs. younger: 89 ± 9, p=0.08), MCHC (older: 33.6 ± 1.4 vs. younger: 34.3 ± 1.8, p=0.05), and WBC (older: 10.2 ± 2.7 vs. younger: 13.1 ± 4.1, p