Faculty Bibliography

Pre-clinical studies in mice and haemophilic dogs have shown that introduction of an adeno-associated viral (AAV) vector encoding blood coagulation factor IX (FIX) into skeletal muscle results in sustained expression of F.IX at levels sufficient to correct the haemophilic phenotype. On the basis of these data and additional pre-clinical studies demonstrating an absence of vector-related toxicity, we initiated a clinical study of intramuscular injection of an AAV vector expressing human F.IX in adults with severe haemophilia B. The study has a dose-escalation design, and all patients have now been enrolled in the initial dose cohort (2 x 10(11) vg/kg). Assessment in the first three patients of safety and gene transfer and expression show no evidence of germline transmission of vector sequences or formation of inhibitory antibodies against F.IX. We found that the vector sequences are present in muscle by PCR and Southern-blot analyses of muscle biopsies and we demonstrated expression of F.IX by immunohistochemistry. We observed modest changes in clinical endpoints including circulating levels of F.IX and frequency of FIX protein infusion. The evidence of gene expression at low doses of vector suggests that dose calculations based on animal data may have overestimated the amount of vector required to achieve therapeutic levels in humans, and that the approach offers the possibility of converting severe haemophilia B to a milder form of the disease

Inhibitors to factor VIII develop in 4-20% of haemophilia A patients, with the percentage rising to 52% in certain subpopulations. The management of inhibitor patients is directed toward stopping acute haemorrhages, providing short-term haemostasis before and after surgery, and inducing immune tolerance to factor VIII (immune tolerance therapy or ITT). Several different protocols have been used for ITT, but they are all centred around ongoing exposure to high doses of factor VIII. High responders (those patients with a large increase in inhibitor level after exposure to factor VIII) are the prime candidates for ITT, but low responders may also benefit from this treatment. It is often necessary to treat bleeding episodes during ITT, because elimination of inhibitors may require many years of therapy. Treatment of haemorrhages in inhibitor patients is reviewed for both low and high responders during ITT and in the absence of ITT. The choice of clotting agent for inhibitor patients who have not yet responded to ITT depends on current and past inhibitor levels, the severity of the haemorrhage, the site of the haemorrhage or the setting in which it occurs (e.g. surgical), and the extent of inhibitor cross-reactivity with porcine factor VIII. Patients with high-titre inhibitors experiencing a critical haemorrhage are generally best managed with bypassing agents (AUTOPLEX T or FEIBA VH), porcine factor VIII or rFVIIa

The majority of children who undergo open-heart surgery with cardiopulmonary bypass (CPB) require perioperative blood transfusion. Blood product requirements are affected by factors such as patient age, underlying cardiac disease, complexity of the surgical procedure, and hemostatic alterations induced by CPB. Transfusion support may include the use of whole blood and/or individual blood components with transfusion practices varying widely based on individual preferences and blood product availability. Approaches to limit allogeneic blood exposure include the use of modified ultrafiltration and smaller bypass circuits, preoperative autologous blood donation and intraoperative blood salvage, and adjunctive antifibrinolytic agents. Potential advantages and disadvantages of the different blood products and pharmacological agents must be considered in managing the pediatric cardiac surgery patient

OBJECTIVE: Resistance to activated protein C (APC) has been identified as a risk factor for thrombotic disease in adults. In over 90% of cases, the basis for the APC resistance is a mutation in the coagulation factor V gene (factor V Leiden) that renders the protein more resistant to inactivation by APC. We sought to determine the prevalence of the factor V Leiden (FVL) mutation in neonates and children who had experienced an arterial or venous thromboembolic event.Study design: We retrospectively analyzed the clinical records of 33 neonates and 52 children with thromboembolic disease. Screening for the FVL mutation was performed by DNA analysis, allowing for identification of patients as normal, heterozygous, or homozygous. RESULTS: Of the 85 patients studied, 12 (14.1%) were heterozygous for FVL; none were homozygous. Of the 47 patients who had arterial central nervous system events, 8 (17%) were positive for the FVL mutation, including 6 of 22 (27%) neonates. Of those patients who had a venous thrombosis, 4 of 32 (12.5%) were FVL positive. None of the 85 patients had protein C deficiency, 3.5% had protein S deficiency, 1.2% had antithrombin III deficiency, and 16.5% had anti-phospholipid antibodies. CONCLUSION: These data suggest that the FVL mutation plays a role in the development of arterial and venous thrombotic events in neonates and children

Patients with homozygous beta-thalassemia are chronically transfused and, if not assiduously chelated, are at risk for cardiac dysfunction. Available data suggest that even in optimally chelated patients, cardiac pathology is abnormal secondary to iron deposition, fibrosis, hypertrophy, and the structural effects of chronic anemia. Evidence of myopericarditis may also be found. Cardiac performance is usually only subtly affected, primarily with diastolic abnormalities not routinely detected on echocardiograms or nuclear scan. In poorly chelated patients, severe heart failure occurs and is easily predictable but invariably fatal, despite treatment with diuretics, vasodilators, inotropes, and antiarrhythmics. Based on successful prevention of heart failure with ACE inhibitors in other forms of cardiomyopathy, we suggest multicenter trials to explore methods to stabilize cardiac function in patients at risk for iron-induced heart disease. Long-term adverse effects of iron deposition, diastolic dysfunction, and abnormal hormone regulation need to be quantitated in patients reaching their third and fourth decades when the potential for ischemic cardiac disease could compound cardiac dysfunction

Controversies about medical practices usually arise from lack of definitive scientific studies. In the presence of continuing controversy about the appropriate hemoglobin level for ventilated (or nonventilated) infants, we can attempt to derive as much useful information as possible. In this article, the authors focus on four subjects: the physiologic role of red cells, the clinical effects of anemia and the proposed clinical benefit of red cell transfusions in preterm infants, the risks associated with transfusions, and the use of recombinant erythropoietin as an alternative to transfusion therapy

BACKGROUND: Hemoglobin (Hb) Bryn Mawr is an unstable Hb variant resulting in congenital hemolytic anemia. This variant Hb also has an increased affinity for oxygen. The perioperative transfusion management of this disorder is described, and the first genomic analysis of this Hb variant is given. CASE REPORT: An 11-year-old boy, heterozygous for Hb Bryn Mawr, was referred for cholecystectomy. Sequence analysis of genomic DNA confirmed that the patients was heterozygous for a T-->C transition in the codon for amino acid 85, causing a substitution of serine for phenylalanine in the beta-globin chain. On the basis of whole-blood O2 dissociation studies, projected tissue O2 delivery would have been suboptimal during general anesthesia; therefore, a partial red cell exchange transfusion was performed to lower variant Hb and prevent tissue hypoxia during surgery. The red cell mass to be exchanged (50%) was determined from the calculated increase in O2 delivery capacity required to maintain an O2 extraction of 4 to 5 mL of O2 per dL of whole blood. The p50 of whole blood from the patients immediately after the exchange transfusion was 16.0 torr. At the time of surgery, the p50 was normal (25.9 torr). The patient's whole blood 2,3 DPG levels were 4.70 mmol per mL of red cells (before transfusion) (normal range = 4.8 +/- 0.3 mmol/mL red cells), 4.07 mmol per mL of red cells (immediately after transfusion), and 4.55 mmol per mL of red cells (48 hours after transfusion). CONCLUSION: This patient with Hb Bryn Mawr was prepared for surgery with a partial exchange transfusion to prevent tissue hypoxia during anesthesia. Decreased 2,3 DPG levels immediately after transfusion resulted in increased O2 affinity of whole blood; however, 48 hours after exchange transfusion, a normal p50 (due to both removal of variant Hb and regeneration of 2,3, DPG) was observed. Partial exchange transfusion is useful in the preoperative management of patients with Hb variants characterized by increased O2 affinity

OBJECTIVE: To determine whether the mean corpuscular hemoglobin concentration (MCHC) or other erythrocyte indexes, as determined by automated cell counters, remains a useful screening test for identifying patients with hereditary spherocytosis (HS). METHODS: Erythrocyte indexes from 112 children with HS who had not undergone splenectomy were compared with those measured in an equal number of healthy, age-matched children. All indexes were derived from measurements obtained by aperture impedance. RESULTS: Mean corpuscular hemoglobin concentration in the HS group was 35.9 gm/dl, significantly higher than in normal control subjects (34.3 gm/dl; p < 0.001). Mean erythrocyte distribution width also was significantly higher in patients with HS (19.3 vs 12.6; p < 0.001). The MCHC distinguishes individuals with HS, with an area under the receiver operating characteristic curve of 0.86. Although not disease specific, an erythrocyte distribution width > 14 has 85% sensitivity and 97% specificity and an area under the receiver operating characteristic curve of 0.92. An MCHC > 35 gm/dl has a sensitivity of 70% and a specificity of 86%. Combining the MCHC and erythrocyte distribution width increases the area under the receiver operating characteristic curve to 0.97. Specificity is 100% and likelihood ratio is infinite when both the MCHC and erythrocyte distribution width are elevated. CONCLUSIONS: The automated MCHC is an effective screening test to identify children with HS. An elevated erythrocyte distribution width adds additional specificity and is itself a powerful screening tool. The combination of the two tests is an excellent predictor for the diagnosis of HS