ALBERT

Description: A 43/4-yr-old black girl with acquired aplastic anemia had an increase in total hemoglobin (Hb) from 4.5 to 16.8 g/dl and fetal hemoglobin (HbF) from 0.8 g/dl (18.8%) to 9.6 g/dl (60.2%) following combined androgen-adrenal steroid therapy. Discontinuation of the drugs was followed by a decline in both HbF and total Hb. Reinstitution of the combined steroids prompted a second rise in total and fetal hemoglobin. During these responses the subject's erythrocytes exhibited an increased i antigen score and a low level of red cell carbonic anhydrase. The glycine:alanine ratio at position 136 of the gamma chains of HbF was of the fetal type (proportion of chains with glycine residues, 0.74). Hemoglobin A2 was low (0.4%). The synthesis of alpha and non-alpha chains was balanced. These results indicate that the stimulation of red cell proliferation in this subject, in response to androgen therapy, resulted in the production of cells with several characteristics of “fetal” erythrocytes.

Description: The advent of therapeutic monoclonal antibodies has enhanced the efficacy of NHL treatment. In recent years, these immuno-therapies have been increasingly used in therapy. We conducted a population-based study of NHL treatment practices in the US using a stratified random sample of patients diagnosed in 1999 with histologically confirmed NHL (n=939) residing in the geographic areas covered by the Surveillance, Epidemiology and End Results program. Blacks and Hispanics were over-sampled to obtain more stable estimates. Patients were followed for vital status through Dec 2001. We performed separate logistic regression analyses to study the potential factors associated with the likelihood of receiving chemotherapy, radiation therapy and the monoclonal antibody, Rituximab. Cox Proportional Hazards regression model was used to study the risk factors associated with survival time. We grouped histological subtypes into five broad categories: B-cell aggressive, B-cell indolent, T-cell generic, cutaneous T-cell, and mantle cell lymphomas. The majority of patients presented with B-cell aggressive or B-cell indolent lymphomas (n=828). Approximately 20% of patients received no therapy. Over 60% of patients received chemotherapy, either alone or in combination. 12% of patients received Rituximab and it was most frequently administered to patients in combination with chemotherapy, especially for patients with B-cell aggressive, B-cell indolent and T-cell generic lymphomas. Only 3% of patients participated in clinical trials. Age and gender were associated with the receipt of chemotherapy: people aged over 75 years, and males were less likely to have received chemotherapy (P=0.01). There were no significant associations between the likelihood of receiving Rituximab and the demographic and clinical factors analyzed. However, our results suggested that African-Americans and people aged over 75 years were less likely to have received immunotherapy. Twenty-four percent of patients received radiation with or without another therapy. When compared to patients with no symptoms at presentation, patients who presented with B-symptoms at diagnosis or those whose B-symptoms were unknown were less likely to have received radiation therapy (OR=0.32 and 0.47 respectively, P=0.0002). Approximately 50% of patients had died by the end of maximum the 3-year follow-up period. Both cause-specific and all-cause mortality was significantly associated with patient age, race/ethnicity, gender, marital status and co-morbid conditions, as well as histological subgroup. Hispanic and Black patients had higher risk of death from both NHL and all-cause (P 75 years, male patients, unmarried patients, or patients with B-symptoms had higher risk of death from either NHL or all-cause (p

Description: A Bangladeshi family is described in which the genes for both hemoglobin E (Hb E) and pyrimidine 5′ nucleotidase deficiency are segregating. An individual homozygous for both these conditions has a severe hemolytic anemia, whereas family members who are homozygous for Hb E are asymptomatic and those homozygous for pyrimidine 5′ nucleotidase deficiency have the mild hemolytic anemia that is characteristic of this disorder. Globin-chain synthesis experiments have shown that the mechanism underlying the interaction between these two genotypes is a marked decrease in the stability of Hb E in pyrimidine 5′ nucleotidase-deficient red blood cells (RBCs). It has also been found that in the enzyme-deficient RBCs in which Hb E is highly unstable, free alpha-chains, though not beta E-chains, acoumulate on the membrane. In view of the increasing evidence that the hemolysis associated with pyrimidine 5′ nucleotidase deficiency results not only from an increase in the level of erythrocyte pyrimidines, but also from inhibition of the hexose monophosphate shunt activity in young erythrocytes, it is likely that the marked instability of Hb E in the enzyme-deficient cells results from oxidant damage acting on a mildly unstable Hb variant. These observations may have important implications for the better understanding of the pathophysiology of Hb E/beta-thalassemia, globally the commonest important form of thalassemia.

Description: The human β-globin gene complex spans a region of 70 kb and contains numerous sequence variants. These variant sites form a 5′ cluster (5′ β-haplotype) and a 3′ cluster (3′ β-haplotype) with strong linkage disequilibrium among the sites within each cluster, but not between the two clusters. The 9-kb region between the 5′ and 3′ clusters has been estimated to have rates of recombination that are 3 to 30 times normal, and the region has therefore been proposed as a ‘hotspot’ of recombination. We describe three families with evidence of meiotic recombination within this ‘hotspot’ of the β-globin gene cluster and in which the cross-over breakpoints have been defined at the sequence level. In one family, the recombination has occurred in the maternal chromosome within a region of 361 bp between positions −911 and −550 5′ to the β-globin gene. In the other two families, the recombination has occurred in the paternal chromosome within a region of approximately 1,100 bp between positions −542 and +568 relative to the β-globin gene cap site. Both regions occur within the 2-kb region of replication initiation (IR) in the β-globin gene domain with no overlap. The IR region contains a consensus sequence for a protein (Pur), which binds preferentially to single-stranded DNA, a role implicated in recombination events.

Description: Hemoglobin E (HbE; 2β226glu-lys), globally the commonest hemoglobin variant, is synthesized at a slightly reduced rate and has a homozygous phenotype similar to heterozygous β thalassemia. Yet, when it is inherited together with a β thalassemia allele, the resulting condition, HbE/β thalassemia, is sometimes characterized by a severe, transfusion-dependent thalassemia major. The severity of this interaction has not been explained. We have explored the possibility that it may reflect the instability of HbE consequent upon globin chain imbalance imposed by the β thalassemia allele. Time-course and pulse-chase globin chain synthesis studies at 37°C on peripheral blood and bone marrow suggest that hemoglobin instability is not significant in steady-state HbE/β thalassemia; this is confirmed by density-gradient centrifugation studies that show no decrease in HbE levels relative to HbA as HbE/β+ thalassemia red blood cells age. Globin binding to membranes was assessed and only  globin chains were found, in contrast to other unstable hemoglobins in which both  and β chains were present. However, in experiments performed on blood from HbE/β thalassemics in the temperature range 39°C to 41°C, there was evidence of instability of HbE, a finding that was also observed in homozygous HbE. These findings suggest that the phenotype of HbE/β thalassemia is primarily the result of the interaction of two β thalassemia alleles; however, hemoglobin instability may be important during febrile episodes, contributing to worsening anemia. © 1998 by The American Society of Hematology.

Description: Analysis of DNA from 852 Island Melanesians has revealed a high frequency of single- and triple-gamma-globin genes in this population. Homozygotes for triple- and single-gamma genes have normal hematologic findings, normal hemoglobin F (HbF) levels, and when there is coexisting alpha thalassemia, appropriate levels of Bart's Hb (gamma 4) at birth. In addition, we have identified an individual with a quadruple-gamma gene chromosome who also has a normal HbF level. All single-gamma genes were A gamma, all triple-gamma genes G gamma G gamma A gamma, and the quadruple-gamma gene G gamma G gamma G gamma A gamma. Analysis of G gamma:A gamma ratios in cord bloods and HbF levels in adults showed that these additional gamma genes are expressed and are down regulated appropriately by the fetal to adult Hb switch. Analysis of the restriction enzyme haplotypes of these various chromosomes indicates that intrachromosomal cross-overs are more likely to have produced these variants than interchromosomal recombination events.

Description: Interleukin 21 (IL-21) is an IL-2 family cytokine produced by activated CD4+ T cells. Potent effects of IL-21 have been observed on the growth, survival, and functional activation of T cells, B cells, and natural killer (NK) cells. A Phase I clinical trial of IL-21 in metastatic melanoma and renal cell carcinoma is currently in progress. We recently reported that IL-21 significantly enhanced rituximab mediated clearance of CD20+ lymphoma cell lines both in vitro and in vivo, and that these effects were potentially mediated through IL-21 enhancement of NK cell capacity to effect antibody dependent cellular cytotoxicity (ADCC). Specifically, NK cells treated with IL-21 showed increased cytotoxicity, granzyme B and IFNg production. Current studies aim to further evaluate the mechanisms by which IL-21 enhances ADCC. A number of observations suggest a multi-factorial basis for IL-21 synergy with rituximab. In a xenograft tumor model, SCID mice were injected IV with HS Sultan cells on day 0. Treatment with recombinant murine IL-21 (mIL-21; starting day 1) combined with rituximab (starting day 3) resulted in significantly increased survival (70% vs. 20% on day 100), compared to rituximab alone. In separate studies, the spleens of mice treated with mIL-21 showed increased numbers of activated macrophages and granulocytes. As macrophages and granulocytes can participate in ADCC, IL-21 synergy with rituximab in vivo may be partly dependent on its activation of these cell types. We have also evaluated whether direct effects of IL-21 on lymphoma cells contribute to enhancement of rituximab efficacy. The xenogeneic B lymphoma models in which IL-21 plus rituximab exhibited enhanced survival are highly aggressive and these models were not shown to respond to treatment with mIL-21 alone. In vitro studies were performed to determine if IL-21 could potentiate the growth inhibitory and pro-apoptotic effects of rituximab. In the absence of effector cells synergistic interaction was not observed. In addition, we tested the ability of IL-21 to enhance cytotoxicity when combined with antibodies targeting non-hematopoietic tumor cells (e.g. trastuzumab). Human NK cells treated with IL-21 displayed significantly increased cytotoxicity in ADCC assays using trastuzumab to target breast cancer cells expressing varying levels of HER-2 antigen. In summary, the current evidence suggests that IL-21 can enhance antibody-mediated tumor cell lysis through activation of multiple effectors of ADCC. Thus IL-21 may prove to be broadly applicable to monoclonal antibody therapy of cancer.

Description: A 12-yr-old Malay boy who was studied because his youngest brother and both his parents had the slow-moving Hb X components (earlier reported to lead to Hb H disease when combined with α-thalassemia) was found to be homozygous for the same slow-moving components. He had splenomegaly and a just palpable liver, mild anemia with microcytosis, hypochromia, slight morphologic changes of the red blood cells, and slight reticulocytosis. Of eight children in the family, six had the trait for the abnormality, one was normal, and one, the propositus, was homozygous. Structural studies of the isolated abnormal hemoglobin showed it to be identical to Hb Constant Spring (Hb CoSp), an α-chain variant with 172 residues instead of the usual 141, the additional 31 being attached to the C-terminal end. In addition to the abnormal α variant for which the propositus was homozygous, he also had normal Hb A and normal Hb A2 with normal α-chains. If the theory that Hb CoSp is due to a structural mutation affecting the terminator codon is correct, this case provides evidence for a duplication of the gene for α-chain production. Results of study of several erythrocyte enzymes are also reported.

Description: The human β-globin gene complex spans a region of 70 kb and contains numerous sequence variants. These variant sites form a 5′ cluster (5′ β-haplotype) and a 3′ cluster (3′ β-haplotype) with strong linkage disequilibrium among the sites within each cluster, but not between the two clusters. The 9-kb region between the 5′ and 3′ clusters has been estimated to have rates of recombination that are 3 to 30 times normal, and the region has therefore been proposed as a ‘hotspot’ of recombination. We describe three families with evidence of meiotic recombination within this ‘hotspot’ of the β-globin gene cluster and in which the cross-over breakpoints have been defined at the sequence level. In one family, the recombination has occurred in the maternal chromosome within a region of 361 bp between positions −911 and −550 5′ to the β-globin gene. In the other two families, the recombination has occurred in the paternal chromosome within a region of approximately 1,100 bp between positions −542 and +568 relative to the β-globin gene cap site. Both regions occur within the 2-kb region of replication initiation (IR) in the β-globin gene domain with no overlap. The IR region contains a consensus sequence for a protein (Pur), which binds preferentially to single-stranded DNA, a role implicated in recombination events.

Description: To try to further define the mechanisms that increase the levels of hemoglobin F (HbF) in the blood of patients with severe forms of β thalassemia, we have studied two comparable populations of hemoglobin E (HbE)/β thalassemics, one regularly transfused and one receiving only occasional blood transfusions. Regular transfusion was associated with a significant decrease in soluble transferrin receptor and erythropoietin levels. Globin chain synthesis studies also show a highly significant decrease in HbF synthesis relative to HbE in the transfused patients. This effect was confirmed by sequential data on one patient, studied before and after the commencement of regular blood transfusion; blood transfusion was followed by a marked increase in the /γ, βE/γ, and HbE/HbF ratios. These data suggest that the high HbF levels in HbE/β thalassemia, and other β thalassemia syndromes, result from increased erythropoietin levels leading to bone marrow expansion, and possibly increased F-cell production, combined with ineffective erythropoiesis giving a survival advantage to F cells. This study also suggests that alteration in blood transfusion regimes must be taken into account when interpreting changes in HbF levels seen in trials of HbF-promoting drugs.