Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product.

Six monoclonal antibodies have been isolated from mice immunized with synthetic peptide immunogens whose sequences are derived from that of the human c-myc gene product. Five of these antibodies precipitate p62c-myc from human cells, and three of these five also recognize the mouse c-myc gene product. None of the antibodies sees the chicken p110gag-myc protein. All six antibodies recognize immunoblotted p62c-myc. These reagents also provide the basis for an immunoblotting assay by which to quantitate p62c-myc in cells.     

Bifunctionality and pseudoisozymes of pyruvate kinase from codfish muscle

Pyruvate kinase has been purified from codfish muscle. The ratio of phosphotransferase and oxalacetate decarboxylase activities remains relatively constant throughout purification steps. These two activities are dependent as well as sensitive to sulfhydryl reagents. In the presence of dithioerythritol, only one molecular form of pyruvate kinase is detected. However, the enzyme exists as four pseudoisozymes in the presence of 2-mercaptoethanol. The pseudoisozymes of codfish pyruvate kinase are interconvertible under the influence of sulfhydryl reagents.     

Structural changes of isolated hepatocytes during treatment with digitonin

The structural changes accompanying digitonin-induced release of enzymes and metabolites from isolated hepatocytes have been studied by scanning and transmission electron microscopy. In the initial phase, characterized by total release of the cytosolic marker enzyme, lactate dehydrogenase, the plasma membrane was immediately damaged, rapidly followed by extensive damage to the endoplasmic reticulum. The shape of the cell, however, was maintained, and the mitochondria and nucleus remained tightly held together by the cytoskeleton. Mitochondria remained intact initially, whereas the cytosol became less electron dense and the nuclear chromatin was more dispersed. An intermediate phase was characterized by total release of adenylate kinase and most of the glucose-6-phosphatase, marker enzymes for the mitochondrial intermembrane space and the endoplasmic reticulum, respectively. The outer mitochondrial membrane was ruptured, but mitochondria maintained their normal matrix electron density. In the final phase, characterized by the beginning of citrate synthase release from the mitochondrial matrix space, the mitochondria became swollen, and only the nucleus, inner and outer mitochondrial membranes, and the cytoskeleton could be clearly distinguished. Although the plasma membrane could not be readily discerned in electron micrographs after the initial phase, the plasma membrane marker enzyme 5'-nucleotidase remained associated with digitonin-treated hepatocytes. Acetyl-CoA carboxylase was released much more slowly than lactate dehydrogenase, indicating some severe restriction on its release. The release of acetyl-CoA carboxylase closely paralleled the release of glucose-6-phosphatase. The controlled exposure of hepatocytes to digitonin, therefore, leads to the sequential release of soluble, compartmentalized cellular components and some membrane-bound components, but the mitochondrial membrane, cytoskeleton and the nucleoskeleton survive even long-term digitonin treatment.     

A phase I trial of recombinant tumor necrosis factor (rTNF) administered by continuous intravenous infusion in patients with disseminated malignancy

rTNF was administered to 28 patients with advanced metastatic cancers by continuous intra venous infusion for 5 consecutive days every 2 weeks. The dose levels were 30, 40, 70, 110, 180 and 290 µg/M²/day. Groups of 3 patients were started at each successive dose level and then on subsequent courses treated with the next dose level through 4 escalations as tolerated. Tumor types were: colon cancer 14; adenocarcinoma of unknown primary, 2; renal cancer, 2; leiomyosarcoma, 2; lung cancer, 1; prostate cancer, 1; thymona, 1; bladder cancer; 1; parotid, 1; Kaposi's sarcoma 2; ovarian 1. Toxicities included fever and chills (usually within the first 8 hours of infusion), fatigue, headache, decreased performance status, hypotension and CNS. All patients experienced leukopenia and thrombocytopenia within 24 hours or less after start of infusion with return of baseline by 72 hours after rTNF was stopped. The fall in these counts averaged 50% and was not dose related. No major changes in liver or renal function, coagulation or blood lipids were seen. Major dose limiting toxicities were fatigue, confusion, thrombocytopenia, seizures, hypotension and decreased performance status. NK cell activity measured against K562 target cells was augmented from about 30% target cell lysis to about 70% target cell lysis over the first 7 days of treatment. Two patients, both with metastatic colon cancer showed transient, objective tumor regression which did not qualify as a partial response. One patient with ovarian cancer had a stable partial response but progressed after 13 courses of treatment. Continuous infusion of TNF can be safely administered to patients with a maximum tolerated dose of only between 30 and 40 µg/M²/day. In addition, the MTD with continuous infusion seems to be highly variable and unpredictable from patient to patient. These data suggest that continuous infusion will not be an optimal way to administer TNF.     

Identification of a common molecular basis for combined 17α-hydroxylase/17,20-lyase deficiency in two Mennonite families

Summary During the course of studies to characterize mutations of the CYP17 gene that cause the 17-hydroxylase-deficient form of congenital adrenal hyperplasia we have discovered two ostensibly unrelated Mennonite families in which affected individuals are homozygous for the same mutation. The defect is a four-base duplication in exon 8 of the CYP17 gene, which alters the reading frame encoding the C-terminal 26 animo acids of cytochrome P450₁₇.     

Effect of distance,aggregation, and habitat on levels of seed predation for two mammal — dispersed neotropical rain forest tree species

The effect of seed aggregation and distance from conspecific trees on seed predation was experimentally examined for two neotropical tree species, Macoubea guianensis (Apocynaceae) and Pouteria sp. (Sapotaceae) in a lowland tropical rain forest in northeastern Peru. Results of these experiments are discussed in the context of the Janzen-Connell model (Janzen 1970; Connell 1971), which predicts decreased seed survival near parent trees due to either density-or distance-responsive mortality, and Howe's model (Howe 1989) which predicts that trees with seeds dispersed in clumps (aggregated) will not suffer density-dependent predation, and will have higher survival of seeds near the parent tree than other trees. We also examined whether predation on seeds of these species was affected by seed placement in or near 30-m-wide strips regenerating after clear-cutting. Both species appeared to be mammal-dispersed but differed in how frugivores handled seeds, seed size, overall fruit crop size, and gemination time. Neither of the two species studied appeared to suffer seed predation in a manner predicted by the Janzen-Connell model, and patterns of seed predation for only one of the species was similar to predictions of Howe's model. For neither species did seed predation along the edge of, or in the center of, regenerating clear cuts differ from predation 15 m into the primary forest. For Pouteria, seed predation in and near regnerating strips was significantly greater than around forest trees, but the opposite pattern held for Macoubea. Overall, seed predation was much greater on Macoubea. The difference in seed predation for these two species was most likely a result of differences in the types of seed predators that attacked these two species.