The gene that is abnormal in the X-linked form of the phagocytic disorder chronic granulomatous disease has been cloned without reference to a specific protein by relying on its chromosomal map position. The transcript of the gene is expressed in the phagocytic lineage of haematopoietic cells and is absent or structurally abnormal in four patients with the disorder. The nucleotide sequence of complementary DNA clones predicts a polypeptide of at least 468 amino acids with no homology to proteins described previously.

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A B S T R A C T The capacity of human phagocytes to generate superoxide anion (02-), a free radical of oxygen, and a possible role for this radical or its derivatives in the killing of phagocytized bacteria were explored using leukocytes from normal individuals and patients with chronic granulomatous disease (CGD). Superoxide dismutase, which removes 02-, consistently inhibited phagocytosis-associated nitroblue tetrazolium (NBT) reduction indicating the involvement of O2 in this process. Similarly, superoxide dismutase inhibited the luminescence that occurs with phagocytosis, implicating 0-in this phenomenon, perhaps through its spontaneous dismutation into singlet oxygen. Subcellular fractions from homogenates of both normal and CGD leukocytes generated 02-effectively in the presence of NADH as substrate. However strating the interrelationship between NBT reduction and 02 generation in phagocytizing cells. Activity of superoxide dismutase, the enzyme responsible for protecting the cell from the damaging effects of 02-, was approximately equal in homogenates of normal and CGD granulocytes. Polyacrylamide electrophoresis separated this activity into a minor band that appeared to be the manganese-containing superoxide dismutase associated with mitochondria and a more concentrated, cyanide-sensitive, cytosol form of the enzyme with electrophoretic mobility that corresponded to that of erythrocyte cuprozinc superoxide dismutase.Superoxide dismutase inhibited the phagocytic killing of Escherichia coli, Staphylococcus aureus, and Streptococcus viridans. A similar inhibitory effect was noted with catalase which removes hydrogen peroxide. Neither enzyme inhibited the ingestion of bacteria. Peroxide and 02-are believed to interact to generate the potent oxidant, hydroxyl radical (-OH). A requirement for -OH in the phagocytic bactericidal event might explain the apparent requirement for both 02-and H202 for such activity. In agreement with this possibility, benzoate and mannitol, scavengers of -OH, inhibited phagocytic bactericidal activity. Generation of singlet oxygen from 02-and -OH also might explain these findings.

The Childrens Cancer Study Group conducted four therapeutic studies on a total of 1006 children with acute nonlymphocytic leukemia from 1972 to 1983. This report describes the therapeutic strategies of these studies and examines trends in induction rates and long-term outcome over this period. The remission induction rate has changed from 58% in 1972 to 1975 to 80% for the period 1980 to 1983, and the induction mortality dropped from 20% to 6%. Four-year survival probabilities from time of diagnosis have almost doubled from 19% to 36%. Few deaths occurred more than 5 years after diagnosis: children surviving in first remission beyond 5 years had a 92% survival rate and an 86% relapse-free survival rate over the next 5 years. In contrast, median survival after a marrow relapse was less than 6 months and the 6-year survival probability was 4%. The leukocyte count was a significant prognostic factor, and although the mortality for infants was high initially, long-term survival was not decreased.

The intact leukocytes of two children with chronic granulomatous disease fail to reduce nitroblue tetrazolium during phagocytosis. This is due to defective operation of an oxidase of reduced nicotinamide adenine dinucleotide that is insensitive to cyanide and that indirectly stimulates the oxidation of glucose-6-phosphate in leukocytes. Such leukocytes undergo no increase in oxygen consumption or in activity of the hexose monophosphate shunt during phagocytosis, although lactate production is normal. The addition of nitroblue tetrazolium to a leukocyte suspension appears to provide a sensitive diagnostic screening test for this disease.

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A B S T R A C T A comparison has been made of the metabolic shifts in human and guinea pig leukocytes when they phagocytize. Respiration of guinea pig polymorphonuclear leukocytes (PMN) and the increment during phagocytosis were each about 2k-fold that of human PMN. This was also true of the direct oxidation of glucose-6-P (hexose monophosphate shunt). Enzymes potentially responsible for these phenomena have been compared in each species. Cyanide-insensitive NADH oxidase and NADPH oxidase were measured and only the former exhibited adequate activity to account for the respiratory stimulus durintg phagocytosis. The hydrogen peroxide formed by this enzyme stimulates the hexose monophosphate shunt by oxidizing glutathione which upon reduction by an NADPH-linked glutathione reductase provides NADP to drive the hexose monophosphate shunt. Other linkages between respiratory stimulation and that of the hexose monophosphate shunt also pertain in the guinea pig.

Because ascorbate potentiates chemotaxis of normal leukocytes, we examined the effect of ascorbate on polymorphonuclear leukocytes from a patient with the Chediak-Higashi syndrome. Chemotactic migration was 104+/-16 leukocytes per 10 fields (mean+/-S.D.) initially and 258+/-44 (P less than 0.001) after ascorbate, as compared to 182+/-10 in controls. There was no bactericidal activity by 40 minutes in the patient's untreated leukocytes. After ascorbate bactericidal activity of patient and untreated control cells was the same. The addition of ascorbate reduced cAMP levels in the patient's cells from a mean of 34.5 pmoles per 10(7) polymorphonuclear leukocytes to 5.9, as compared to a control value of 3.1+/-1.4. The association of elevated cAMP and impaired function in the polymorphonuclear leukocytes of patients with the Chediak-Higashi syndrome may be related to abnormal microtubular assembly.