A specific and potent immunotoxin composed of antibody ZME-018 and the plant toxin gelonin

Murine monoclonal antibody ZME-018 recognizes a 240 Kda glycoprotein present on the surface of most human melanoma cells and on over 80% of human biopsy specimens tested. Gelonin is a ribosome-inactivating plant toxin similar in nature and rivaling the activity of ricin A chain. ZME-018 was coupled to purified gelonin using the reagents SPDP and 2-iminothiolane. The ZME-gelonin conjugate was purified by S-300 Sephacryl and Blue Sepharose chromatography, removing unreacted gelonin and antibody, respectively. PAGE analysis showed that ZME was coupled to 1, 2, or 3 gelonin molecules. The ZME-gelonin conjugate was 10(6)-fold more active than gelonin itself in inhibiting the growth of log-phase human melanoma cells in culture. The immunoconjugate was not cytotoxic to antigen negative T-24 (human bladder carcinoma) cells. Treatment of melanoma cells with recombinant IFN-alpha or TNF substantially augmented the cytotoxicity of the immunoconjugate while treatment with IFN-gamma had a minor effect. Using the human tumor colony assay of melanoma cells obtained from fresh biopsy specimens, greater than 90% growth suppression was observed in 2 of 4 samples tested at a concentration of 250 ng/ml. In addition, 25% growth suppression was observed with a third sample tested, and no growth suppression was observed in 1 sample. Thus, clonogenic melanoma cells are sensitive in vitro to the cytotoxic activity of this immunotoxin at concentrations which we presume are pharmacologically relevant.     

PAS IV, an integral membrane protein of mammary epithelial cells, is related to platelet and endothelial cell CD36 (GP IV).

PAS IV is a 78-kDa (bovine) to 80-kDa (human) integral membrane glycoprotein of unknown function which is found in mammary epithelial cells. We now report the purification of human PAS IV and native bovine PAS IV from the milk fat globule membrane (MFGM), a preparation of apical plasmalemma from epithelial cells of lactating mammary tissue. N-Terminal sequence analyses of human and bovine PAS IV revealed homology to the N-terminal sequence of the 88-kDa human endothelial and platelet glycoprotein CD36. The similarity of MFGM PAS IV to platelet CD36 was further established by immunoblots of purified platelet CD36 and MFGM PAS IV with MFGM PAS IV specific antiserum. The removal of N-linked oligosaccharides from PAS IV and CD36 by treatment with endoglycosidase F reduced the apparent Mr of both proteins to approximately 57,000. These data suggest that PAS IV and CD36 are similar if not identical polypeptides that undergo cell type specific glycosylation.     

The expression of granulocyte/macrophage colony-stimulating factor in activated mouse lymphocytes declines with age

The expression of granulocyte/macrophage colony-stimulating factor (GM-CSF) was studied in spleen lymphocytes isolated from male C57BL/6J mice of 6, 20, and 29 months of age. GM-CSF expression (biological activity and mRNA level) was maximum after culturing the lymphocytes for 45 hr with concanavalin A and phorbol myristate acetate. The induction of both GM-CSF activity and mRNA levels was observed to decline over 60% between 6 and 29 months of age. The age-related decline in the level of GM-CSF paralleled the age-related decline in the mRNA levels of interleukin-2 and interleukin-3.     

Obstetrics and Gynecology: Considerations in Career Selection

Current training programs in obstetrics and gynecology are not producing an excess of specialists in view of future manpower needs. In addition to being specialists and consultants, obstetrician-gynecologists also function as providers of primary care for women. During the last decade, three formal sub-specialties of obstetrics and gynecology have evolved: gynecologic oncology, maternal-fetal medicine and reproductive endocrinology. These have improved patient care and have altered the structure of resident education. With more American medical school graduates entering this specialty, the quality of resident applicants has improved, creating intense competition for desirable training positions. Those inclined toward a career in obstetrics and gynecology can be assured that it will provide an increasingly favorable and challenging environment for professional activity in the future.     

Lethal effect of polymyxin B sulfate in experimental Proteus rettgeri infection in mice

The mouse model of intraperitoneal sepsis with Proteus rettgeri was used to evaluate the anti-endotoxic effect of polymyxin B sulfate. An unexpected reversal of the usual protective effect of polymyxin in experimental enterobacterial sepsis was observed in which the lethality of the infection was enhanced.     

The reversal of inhibition of protein synthesis by double-stranded RNA in lysed rabbit reticulocytes with fructose 6-phosphate.

Fructose 6-phosphate (1.4 mM – 3.0 mM) effectively prevents the inhibition of protein synthesis in unfractionated rabbit reticulocyte lysates by the presence of double-stranded RNA (poly rI:poly rC, 1 μg/ml). Glucose 6-phosphate, but not fructose 1,6-diphosphate, is equally as effective as fructose 6-phosphate. The data suggest that fructose 6-phosphate prevents the formation of a protein synthesis inhibitor induced by double-stranded RNA.     

Reaction of phenylglyoxal with arginine. The effect of buffers and pH.

Phenylglyoxal reacts much more rapidly with N²-acetylarginine than with either N²-acetyllysine or N-acetylcysteine. The rate of the reaction of phenylglyoxal with either N-acetylarginine or arginine increases with increasing pH from 7.5 to 11.5. The model reaction with arginine is much faster in bicarbonate, diethylamine, or triethylamine buffer than in N-ethylmorpholine, borate, phosphate, or Tris buffer. This activation by various buffers should be taken into consideration when glyoxal derivatives are used to modify arginyl residues.     

Purification and properties of the hemagglutinin from Wistaria floribunda seeds.

Extracts of Wistaria floribunda seeds contain separable erythroagglutinating and lymphocyte mitogenic activities. We wish to report the purification and characterization of the erythroagglutinating lectin of these seeds. A phosphate-buffered saline (PBS) extract of the ground seeds was made to 50% ethanol and the precipitate, which contained both the agglutinin and mitogen, was dissolved in PBS. The erythroagglutinating activity was adsorbed onto insoluble polyleycyl derivatized A + H active hog gastric mucin. After desorption with 0.2 M D-galactose and removal of the sugar by dialysis, the eluate displayed three protein bands on polyacrylamide gel electrophoresis. The major component represented 85% of the mixture. Immunoelectrophoresis of the mixture demonstrated immunochemical identity among the proteins. Gel filtration through Sephadex G-200 resulted in purification of the major component. Based upon the composition and subunit molecular weight, it was concluded that the three components represent a dimer, tetramer, and octamer of a single glycopolypeptide chain of 28 000. The erythroagglutinin has a pI at pH 5.4 and one cystine per dimeric unit.