A life-long quest to understand and treat genetic blood disorders.

This year's Lasker-Koshland Special Achievement Award in Medical Science is conferred on Sir David Weatherall for his 50 years of dedication to biomedical research, his groundbreaking discoveries about genetic blood diseases, and his life-long passion for bringing improved medical care to the developing world.

The Genetic Landscape of Diamond-Blackfan Anemia.

Diamond-Blackfan anemia (DBA) is a rare bone marrow failure disorder that affects 7 out of 1,000,000 live births and has been associated with mutations in components of the ribosome. In order to characterize the genetic landscape of this heterogeneous disorder, we recruited a cohort of 472 individuals with a clinical diagnosis of DBA and performed whole-exome sequencing (WES). We identified relevant rare and predicted damaging mutations for 78% of individuals. The majority of mutations were singletons, absent from population databases, predicted to cause loss of function, and located in 1 of 19 previously reported ribosomal protein (RP)-encoding genes. Using exon coverage estimates, we identified and validated 31 deletions in RP genes. We also observed an enrichment for extended splice site mutations and validated their diverse effects using RNA sequencing in cell lines obtained from individuals with DBA. Leveraging the size of our cohort, we observed robust genotype-phenotype associations with congenital abnormalities and treatment outcomes. We further identified rare mutations in seven previously unreported RP genes that may cause DBA, as well as several distinct disorders that appear to phenocopy DBA, including nine individuals with biallelic CECR1 mutations that result in deficiency of ADA2. However, no new genes were identified at exome-wide significance, suggesting that there are no unidentified genes containing mutations readily identified by WES that explain >5% of DBA-affected case subjects. Overall, this report should inform not only clinical practice for DBA-affected individuals, but also the design and analysis of rare variant studies for heterogeneous Mendelian disorders.

Cholesterol: the debate should be terminated.

Here, I offer personal perspectives on cholesterol homeostasis that reflect my belief that certain aspects of the debate have been overstated.-Nathan, D. G. Cholesterol: the debate should be terminated.

Hemoglobin disorders: a look to the future.

In this issue of Blood, Locatelli et al compare the results of histocompatible family donor bone marrow and cord blood transplants (BMT and CBT) for severe ß thalassemia (SBT) and sickle cell disease (SCD) as experienced by the Eurocord and European Blood and Marrow Transplantation group and collaborating centers in the United States, Hong Kong, and Israel between 1994 and 2005. Obviously, many changes in medical care and particularly MHC typing occurred over that decade, so this retrospective represents a moving target, but some firm points can be made for which we are indebted to this excellent group.

Sickle cell vaso-occlusion causes activation of iNKT cells that is decreased by the adenosine A2A receptor agonist regadenoson.

Adenosine A2A receptor (A2AR) agonists reduce invariant natural killer T (iNKT) cell activation and decrease inflammation in sickle cell disease (SCD) mice. We conducted a phase 1 trial of the A2AR agonist regadenoson in adults with SCD. The target dose was 1.44 µg/kg/h. iNKT cell activation was evaluated using antibodies targeting the p65 subunit of nuclear factor-κB (phospho-NF-κB p65), interferon-γ (IFN-γ), and A2AR. Regadenoson was administered to 27 adults with SCD. We examined 21 patients at steady state and 6 during painful vaso-occlusive crises (pVOC). iNKT cell activation was also measured in 14 African-American controls. During pVOC, the fraction of iNKT cells demonstrating increased phospho-NF-κB p65 and A2AR expression was significantly higher compared with controls (P < .01) and steady-state patients (P < .05). IFN-γ expression was also significantly higher compared with controls (P = .02). After a 24-hour infusion of regadenoson during pVOC, phospho-NF-κB p65 activation in iNKT cells decreased compared to baseline by a median of 48% (P = .03) to levels similar to controls and steady-state SCD. No toxicities were identified. Infusional regadenoson administered to adults with SCD at 1.44 µg/kg/h during pVOC decreases activation of iNKT cells without toxicity.

Advances in sickle cell therapies in the hydroxyurea era.

In the hydroxyurea era, insights into mechanisms downstream of erythrocyte sickling have led to new therapeutic approaches for patients with sickle cell disease (SCD). Therapies have been developed that target vascular adhesion, inflammation and hemolysis, including innovative biologics directed against P-selectin and invariant natural killer T cells. Advances in hematopoietic stem cell transplant and gene therapy may also provide more opportunities for cures in the near future. Several clinical studies are underway to determine the safety and efficacy of these new treatments. Novel approaches to treat SCD are desperately needed, since current therapies are limited and rates of morbidity and mortality remain high.

Fetal hemoglobin levels and morbidity in untransfused patients with ß-thalassemia intermedia.

To evaluate the association between fetal hemoglobin (HbF) levels and morbidity in ß-thalassemia intermedia (TI), we analyzed data from 63 untransfused patients who had also never received HbF induction therapy. Patient records were reviewed for any history of 10 predefined morbidities. Laboratory measurements for markers of ineffective erythropoiesis were also obtained. The mean age of patients was 32.1 years, 47.6% were males, and the median HbF level was 37.2%. HbF levels correlated positively with total hemoglobin, yet negatively with growth differentiation factor-15 and non-transferrin-bound iron levels. Median HbF levels were significantly lower in patients with the majority of evaluated morbidities than in those without. There was a strong negative adjusted linear correlation between the HbF level and the total number of morbidities (R(2) = 0.825, P < .001). The HbF threshold of 63.7% had 95.5% sensitivity and 100% specificity for ensuring absence of morbidity. There exists a strong association between HbF levels and morbidity in the subset of untransfused patients with TI.

Pulmonary hypertension and nitric oxide depletion in sickle cell disease.

During the past decade a large body of experimental and clinical studies has focused on the hypothesis that nitric oxide (NO) depletion by plasma hemoglobin in the microcirculation plays a central role in the pathogenesis of many manifestations of sickle cell disease (SCD), particularly pulmonary hypertension. We have carefully examined those studies and believe that the conclusions drawn from them are not adequately supported by the data. We agree that NO depletion may well play a role in the pathophysiology of other hemolytic states such as paroxysmal nocturnal hemoglobinuria, in which plasma hemoglobin concentrations are often at least an order of magnitude greater than in SCD. Accordingly, we conclude that clinical trials in SCD designed to increase the bioavailability of NO or association studies in which SCD clinical manifestations are related to plasma hemoglobin via its surrogates should be viewed with caution.

Sickle cell disease (SCD), iNKT cells, and regadenoson infusion.

A humanized murine sickle cell-disease (SCD) model (NY1DD) has been used to study ischemia/reperfusion injury (IRI) in sickle cell anemia, and iNKT cells (a very small subset of murine and human T cells) have been found to instigate such injury in this model. Furthermore, levels of activated iNKT cells are generally elevated in the circulation of patients with SCD. Because activated iNKT cells are rich in adenosine A2A receptors which, when agonized, down-regulate the inflammatory cytokine expression that characterizes the cells, we have conducted a phase 1 trial of a constant infusion of low-dose regadenoson (an adenosine analogue with high A2A receptor specificity) to determine its safety and the capacity of a safe dose to down-regulate circulating iNKT cells in patients with SCD. We have found two dose rates that are both safe and effective and now plan a controlled Phase 2B clinical trial to determine whether our highest dose, administered as a 48-hour constant infusion, will induce faster remission in both painful vaso-occlusive crisis (pVOC) and acute chest syndrome (ACS).

Targeting iNKT cells for the treatment of sickle cell disease.

Sickle cell disease (SCD) causes widely disseminated vaso-occlusive episodes. Building on evidence implicating invariant NKT (iNKT) cells in the pathogenesis of ischemia/reperfusion injury, recent studies demonstrate that blockade of iNKT cell activation in mice with SCD reduces pulmonary inflammation and injury. In patients with SCD, iNKT cells in blood are increased in absolute number and activated in comparison to healthy controls. iNKT cell activation is reduced by agonists of adenosine 2A receptors (A(2A)Rs) such as the clinically approved coronary vasodilator, regadenoson. An ongoing multi-center, dose-finding and safety trial of infused regadenoson, has been initiated and is providing preliminary data about its safety and efficacy to treat SCD. Very high accumulation of adenosine may have deleterious effects in SCD through activation of adenosine 2B receptors that are insensitive to regadenoson. Future possible therapeutic approaches for treating SCD include selective A(2B)R antagonists and antibodies that deplete iNKT cells.

Sickle cell disease and stroke.

Twenty-four percent of sickle cell disease (SCD) patients have a stroke by the age of 45 years. Blood transfusions decrease stroke risk in patients deemed high risk by transcranial Doppler. However, transcranial Doppler has poor specificity, and transfusions are limited by alloimmunization and iron overload. Transfusion withdrawal may be associated with an increased rebound stroke risk. Extended blood typing decreases alloimmunization in SCD but is not universally adopted. Transfusions for thalassemia begun in early childhood are associated with lower rates of alloimmunization than are seen in SCD, suggesting immune tolerance. Optimal oxygen transport efficiency occurs at a relatively low hematocrit for SCD patients because of hyperviscosity. Consequently, exchange rather than simple transfusions are more effective in improving oxygen transport efficiency, but the former are technically more demanding and require more blood units. Although viscosity is of importance in the noncerebral manifestations of SCD, inflammation may play a larger role than viscosity in the development of large-vessel stroke. The future of SCD stroke management lies in the avoidance of transfusion. Hydroxyurea and anti-inflammatory measures may reduce the need for transfusion. Recent genome-wide association studies may provide methods for modulating fetal hemoglobin production enough to attenuate stroke risk and other complications of SCD.

The several Cs of translational clinical research.

Perhaps because I am a veteran of the "good old days" (they were really quite bad), young physicians who hope to become clinical investigators often ask me how they might establish their careers. Many are more than a little worried about their futures and often have trouble envisioning a career path that is financially secure for themselves and their families. The grumbling of clinical investigators a few years their senior enhances their angst. So I try to encourage these young physicians because I know the great intellectual (if not monetary) rewards of the field and because I know that the future of medicine absolutely depends on clinical investigators. The following is what I try to say to them.

The Pathophysiology of Acquired Aplastic Anemia: Current Concepts Revisited.

Idiopathic acquired aplastic anemia is a rare, life-threatening bone marrow failure syndrome characterized by cytopenias and hypocellular bone marrow. The pathophysiology is unknown; the most favored model is of a dysregulated immune system leading to autoreactive T-cell destruction of hematopoietic stem and progenitor cells in a genetically susceptible host. The authors review the literature and propose that the major driver of acquired aplastic anemia is a combination of hematopoietic stem and progenitor cells intrinsic defects and an inappropriately activated immune response in the setting of a viral infection. Alterations in bone marrow microenvironment may also contribute to the disease process.