Molecular genetic studies in black families with sickle cell anemia and unusually high levels of fetal hemoglobin.

Clinical, hematologic, and molecular genetic studies are reported for five families with SS patients having unusually high fetal hemoglobin (Hb F) levels (mean 28.3%, range 19-42%). Some of the individuals were symptom-free and one was not anemic. However, some were symptomatic despite a very high Hb F. Neither the Hb F level nor the F cell distribution entirely explained the variation in clinical severity. Molecular genetic studies identified the Senegal haplotype with the associated -158 G gamma (C----T) mutation in two of the five families. The -202 G gamma (C----G) mutation was not found in any of the individuals studied. Sequencing of the gamma-globin gene promoters to detect genetic high F determinants not detectable by restriction digestion was not performed. All AS parents and AS siblings demonstrated elevated F cells when the Senegal/-158 G gamma (C----T) mutation was present with either the beta S or beta A allele. Double heterozygosity for two different high F determinants in some SS patients is suggested by the studies in at least one family. Discordance among siblings in clinical and hematologic manifestations in two families provides additional evidence for loci regulating Hb F cell production which are not linked to the beta-globin gene clusters.

Newborn screening for hemoglobinopathies in Colorado. The first 10 years.

In Colorado, newborn screening for hemoglobinopathies by cellulose acetate and citrate agar electrophoresis of dried capillary blood spots was established in 1979. We reviewed the results of screening 528,711 infants through 1988. Forty-seven infants with sickle cell diseases and 27 infants with other hemoglobin diseases were identified. The initial screening failed to detect sickle cell anemia in 4 infants, but the hemoglobinopathy in 3 of these infants was diagnosed correctly by routine retesting of those with suspected sickle cell trait. A total of 47 infants with sickle cell diseases were followed through September 1989. There was no mortality among these infants. The screening test identified 3779 infants (1:140 births) with a suspected hemoglobin trait; confirmatory retesting was obtained in 53%. The results of our experience confirm the value of newborn screening for hemoglobinopathies but suggest that a more sensitive test would improve the program.

Normal vitamin E status in sickle hemoglobinopathies in Colorado.

Because previous studies of serum or plasma vitamin E (E) levels reported a high prevalence of E deficiency in patients with sickle cell anemia (SCA), we studied the E status in 101 patients with SCA in Colorado using both levels of serum E and ratios of serum E to total lipid (E:L). Compared with age-, sex-, and race-matched controls, 1 of 70 patients with homozygous SCA (SS), 1 of 7 with sickle beta+-thalassemia, and 0 of 24 with hemoglobin SC disease had E deficiency according to E:L and all were E-sufficient based on serum E levels. Serum cholesterol levels, lower in SS patients than in control subjects, correlated more strongly with serum E levels than did total serum lipid levels in control subjects and SS patients; hence, the ratio of serum E to cholesterol may be a useful indicator of E status in these patients. We conclude that vitamin E deficiency rarely occurs in SCA patients in Colorado.

Intranasal administration of deferoxamine to iron overloaded patients.

We examined the effect of intranasal administration of deferoxamine on iron excretion in seven patients with iron overload secondary to chronic transfusion therapy. Deferoxamine was administered in doses of 0.75 to 3.0 gm given over 12 hours in a variety of dosing schedules. There was a probable, though not significant, dose response relationship between the amount of iron excreted and the dose administered. The amount of iron excreted was 10%-15% of that obtained using the same dosage of deferoxamine given by the subcutaneous route over the same time period. Hourly administration was more effective than less frequent administration. Addition of taurodeoxycholate to deferoxamine did not increase its absorption as measured by the levels of iron excretion. Side effects were few and consisted mainly of mild nasal irritation and a bad taste in the mouth. Nasal administration of deferoxamine may be a useful adjunct to iron chelation in patients receiving chronic transfusion therapy, particularly in those who are noncompliant with parenteral means of administration.

Gallbladder function is altered in sickle hemoglobinopathy.

Despite comparable rates of hemolysis, only 50% of patients with sickle hemoglobinopathy (SH) develop pigment gallstones by age 20 yr. Thus, pathogenetic factors, other than hemolysis, may contribute to gallstone formation. In the present study we determined whether gallbladder function, measured by real-time ultrasonography or bile acid metabolism, determined by isotope dilution-mass spectrometry, were altered in adolescents and young adults with SH. Compared with healthy controls, SH subjects had larger fasting (27 +/- 16 vs. 15 +/- 5 ml, p less than 0.02), and residual (8 +/- 6 vs. 4 +/- 2 ml, p less than 0.03) volumes of the gallbladder, but similar rates of emptying (0.029 +/- 0.016 vs. 0.034 +/- 0.029 min-1) and percentage of fasting volume emptied (71% +/- 13% vs. 72% +/- 14%). In SH subjects, the volume and emptying of the gallbladder were similar between those with and without gallstones. Some SH subjects had stasis of bile within the gallbladder, as demonstrated by isotopic disequilibrium between the circulating bile acid pool and bile stored in the gallbladder. Subjects with SH with gallstones tended to have smaller bile acid pools than SH subjects without gallstones (81 +/- 11 vs 163 +/- 91 mumol/kg, p = 0.051). We conclude that adolescents and young adults with SH have enlarged gallbladders that retain an increased postprandial volume of bile. Bile retention within the gallbladder may lead to stasis and contribute to the pathogenesis of pigment gallstones.

Zinc status in children and young adults with sickle cell disease.

Previous studies have suggested an association of zinc deficiency and short stature in some children and adults with sickle cell disease (SCD). As a result, zinc supplementation has been recommended for these patients. The mechanism for zinc deficiency in certain patients with SCD is unknown, although renal loss of zinc has been suggested as a contributing factor. The zinc status of 29 subjects with SCD and 18 black controls was studied. No evidence of zinc deficiency in our population with SCD was found when plasma and cellular zinc levels were measured. Likewise, levels of two zinc-dependent enzymes, alkaline phosphatase and delta-aminolevulinic acid dehydratase, were normal in these subjects with SCD. Although adolescent subjects with SCD tended to be shorter than control subjects, there was no correlation between the height-forage z score and plasma zinc levels (r = -.31). It was concluded that zinc deficiency was not present in our population with SCD, and that there was no correlation between plasma zinc levels and the height-for-age z score in growing adolescent patients with SCD. These findings suggested that zinc supplementation may not be necessary in all patients with SCD.

Experience with donors matched for minor blood group antigens in patients with sickle cell anemia who are receiving chronic transfusion therapy.

Clinically significant alloimmunization to red cell antigens occurred in 28 percent of transfused patients in a sickle cell clinic. Therefore, a prospective study was undertaken to determine whether matching donors carefully for 17 blood group antigens would diminish the risk of further alloimmunization in patients on a chronic transfusion program. Alloantibodies had developed previously in 8 of the 12 patients. After chronic transfusion with selected donors, four new antibodies developed in three patients. Three antibodies were due to errors in phenotyping or matching, and one was due to an antigen that was not tested for in the protocol. The incidence of developing antibodies per unit transfused was diminished tenfold when selected donors were used. Autoantibodies developed in five patients (42%), but these did not seriously interfere with the transfusion therapy. It was concluded that matching for red cell antigens may diminish the incidence of alloimmunization in patients with sickle cell anemia requiring transfusion.

Splenic syndrome at mountain altitudes in sickle cell trait. Its occurrence in nonblack persons.

Six consecutive cases of splenic syndrome at mountain altitudes in persons with sickle cell trait are reported and the literature is reviewed. All six cases occurred in men who experienced the acute onset of severe left-upper-quadrant abdominal pain within 48 hours of arrival in Colorado from lower altitudes. All six patients were phenotypically nonblack. Three patients experienced their symptoms at moderate altitudes of 1,609 to 2,134 m (5,280 to 7,000 ft) above sea level. All recovered with medical management and none required splenectomy, although functional hyposplenia was a sequela in at least one patient. The possibility that nonblack persons with sickle cell trait may be at greater risk than black persons with sickle cell trait for the development of splenic syndrome at moderate altitude is discussed.

Intermittent combination chemotherapy with or without bacillus Calmette-Guérin for treatment of acute lymphoblastic leukemia of childhood.

Seventy-four children ranging in age from 6 months to 17.5 years with acute lymphoblastic leukemia newly diagnosed between 1976 and 1979 were entered on a study incorporating intermittent chemotherapy with or without the addition of bacillus Calmette-Guérin (BCG). The chemotherapy program consisted of induction with vincristine, dexamethasone, and intrathecal methotrexate, intensification with adriamycin and asparaginase, central nervous system treatment with cranial irradiation and intrathecal methotrexate, and continuation treatment with 5-day courses of combination chemotherapy administered every three weeks. The first phase of continuation therapy incorporated vincristine, adriamycin, 6-mercaptopurine, and dexamethasone. In the second phase, oral methotrexate was substituted for the adriamycin in non-T-cell patients; in T-cell patients, cytosine arabinoside or cyclophosphamide and methotrexate in alternating cycles were substituted for the adriamycin and asparaginase was added. Total duration of therapy was approximately 2.5 years. Connaught BCG was administered by Heaf gun on days 8 and 15 of each 3-week cycle for the first 8 months of treatment in approximately one-third of the patients. Actuarial disease-free survival with a median follow-up of 59 months shows no difference in outcome between the BCG and non-BCG poor-risk patients. However, there is an improvement in disease-free survival of BCG-treated good- and average-risk girls (P = 0.04). While patients were actively receiving BCG there was also a trend toward the development of fewer significant infections than when patients were not receiving BCG (P = 0.85). Toxicities from BCG administration included satellite rashes, local tenderness, lymphadenopathy, secondary infection, and residual scars. Overall disease-free survival by actuarial analysis is 60% at 6 years; for patients with unfavorable prognostic features it is 40%. In this trial the addition of BCG prolonged the disease-free survival of girls with good- and average-risk prognostic features and also may have decreased the susceptibility to infection while it was being administered. However, the benefit does not appear sufficient to warrant its routine use, especially in view of the toxicities encountered.

Probable graft-vs-graft reaction in an infant after exchange transfusion and marrow transplantation.

A newborn with graft-vs-host (GVH) disease following an exchange transfusion was treated by attempting to eradicate the incompatible graft and to reconstitute the child hematologically and immunologically with a bone marrow transplant. The patient was a female term infant (blood group B, Rh+ Coombs test positive) who received a one-unit group O, Rh- exchange transfusion from an unrelated female donor for hyperbilirubinemia due to ABO incompatibility on day 2. Signs of acute GVH disease began on day 8 and the clinical diagnosis was supported by skin biopsy. With antithymocyte globulin and high dose dexamethasone, the GVH reaction improved somewhat. Cyclophosphamide, 200 mg/kg total dose, was given over four days followed by a marrow graft from a brother who was HLA-A, B identical, and probably also D locus compatible in mixed lymphocyte culture. All signs of GVH resolved with cyclophosphamide treatment and hematologic reconstitution was evident by 14 days after transplant. Two weeks later the GVH reaction and aplastic anemia recurred and Y chromatin was detected in only 6% of marrow cells. The infant died on day 80. Autopsy showed disseminated candidiasis, disseminated cytomegalovirus infection, thymic dysplasia, hypoplastic marrow, and other histopathologic changes consistent with GVH disease. The persistence of female cells in blood and bone marrow and the destruction of the reconstituted marrow suggest that the original incompatible transfusion-derived graft was not eliminated and that it ultimately rejected the histocompatible marrow graft.

Effect of subcutaneous deferoxamine and oral vitamin C on iron excretion in congenital hypoplastic anemia and refractory anemia associated with the 5q-syndrome.

Chronic refractory anemia associated with congenital hypoplastic anemia (CHA, Blackfan-Diamond syndrome) and with the 5q-syndrome may require chronic transfusion therapy to sustain life. Hemosiderosis and death from chronic iron overload may result from such a program. The effect of subcutaneous (SC) deferoxamine (DF) and supplemental oral vitamin C (vit. C) on urinary iron excretion was studied in two patients with congenital hypoplastic anemia and one patient with 5q-syndrome. In the two patients with CHA, urinary iron excretion in response to DF given SC over 24 hours was comparable to the results following intravenous (I.V.) administration. Both of these cases had low levels of plasma ascorbate on initial evaluation and excreted more iron in response to two different doses of DF after they had received supplemental vit C and their stores were repleted. Significant iron excretion occurred in all three patients for 12 hours during the SC infusion of DF and for 12 hours after the end of the infusion. In all three patients, increasing the dose of DF up to 3-4 g given SC over 12 hours resulted in a linear increase in iron excretion. Once normal body stores of ascorbate were achieved by oral supplementation, increasing doses of vit C did not appear to cause a further increment in iron excretion. DF administered by a slow SC infusion appears to be an effective approach to iron overload in patients with refractory anemia and hemosiderosis secondary to chronic transfusions. Only small amounts of supplemental vit. C necessary to sustain adequate body stores are required for optimal iron excretion.

Regression on oxymetholone-induced hepatic tumors after bone marrow transplantation in aplastic anemia.

Treatment of acquired aplastic anemia with androgens has been occasionally associated with the development of hepatic tumors. We have studied a 13-year-old boy with idiopathic aplastic anemia in whom oxymetholone treatment was associated with a partial hematological remission. Thirty-four months later, however, the patient developed multiple hepatic tumors. When oxymetholone therapy was discontinued, the aplastic anemia relapsed. He then underwent bone marrow transplantation from his HLA-A, B, and D-compatible sibling. This was followed by hematological and immunological reconstitution. The hepatic tumors underwent progressive regression after bone marrow transplantation. The patient is now 3 years post-bone marrow transplantation and is in complete remission of his aplastic anemia with no evidence of detectable liver tumors.

Sickling crises and altitude. Occurrence in the Colorado patient population.

Seventy-five black patients with sickle hemoglobinopathies who are followed by the Colorado Sickle Cell Treatment and Research Center, and 172 of their family members were evaluated by retrospective interview for the occurrence of sickling crises when traveling in the Rocky Mountains or by aircraft. Twenty per cent of 39 patients with sickle cell anemia (Hgb SS) (Hgbs S/C and S/T) have developed crises when traveling in the mountains above 2000m. Vaso-occlusive crises predominated in the SS group and splenic crises occurred primarily in those with Hgbs S/C and S/T. Approximately 20 per cent of those with S/C and S/T, but none with SS, had crises when flying in pressurized aircraft. Among 103 family members with sickle cell trait (Hgb AS), no significant risk of developing crises could be identified with either mountain or pressurized aircraft travel.

Remission of relapsed leukaemia during a graft-versus-host reaction. A "graft-versus-leukaemia reaction" in man?

Acute lymphoblastic leukaemic in two boys relapsed after engraftment of marrow from siblings identical at HLA A, B, and D loci but went into remission during subsequent graft-versus-host reactions without specific anti-leukaemia therapy. Later leukaemic relapse was primarily in extramedullary sites, with little or no involvement of bone-marrow, liver, or spleen. Cytogenetic studies in both cases showed that the relapsed leukaemic blasts were those of the recipients while marrow cells and blood lymphocytes detected during marrow remission originated from the female donors. Blood lymphocytes from one of the recepients kiled. 51Cr-labelled autologous lymphoblast. The prolonged bone-marrow remission in the face of active and even massive extramedullary leukaemia suggests a graft-versus-leukaemia reaction in these two patients.