Anti-Melanoma Monoclonal Antibody 225-28S: Evaluation of Toxicity in Man

To investigate possible undesirable effects due to the intravenous administration of a reagent of a xenogenic nature (monoclonal antibody 225-28S) in man, a toxicologic study was carried out on 85 patients with metastatic cutaneous melanoma. Two reagents were tested in this study: purified monoclonal antibody (MoAb) 225-28S and its F(ab')2 fragment. Purified MoAb was labelled with 131I and F(ab')2 fragment with 131I, or 123I, or 111In or 99Tc. The quantity of MoAb or F(ab')2 injected ranged from 14 to 750 μg, and the specific activity from 37.0 to 2116.4 MBq/mg. The total radioactivity injected varied from 25.9 to 891.7 MBq/mg. In addition to a careful clinical examination, the following tests were done to monitor possible adverse effects: blood glucose, azotemia, RBC, WBC, platelet count, serum creatinine, creatinine clearance, plasma electrolyte levels, serum proteins, albumin/globulin ratio, serum bilirubin, SGOT, SGPT, γGT, and CPK. These tests were done before the injection and on days 7 and 14. No patient experienced adverse general effects like fever, nausea, vomiting or allergic reactions. None of the aforementioned hematometric and biochemical tests showed significant variations compared with the initial values. It is concluded that a single injection of these reagents at the dosages tested is completely atoxic.

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Conformation-Specific Inhibitory Anti-MMP-7 Monoclonal Antibody Sensitizes Pancreatic Ductal Adenocarcinoma Cells to Chemotherapeutic Cell Kill

Cancers ◽

10.3390/cancers13071679 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1679

Author(s):

Vishnu Mohan ◽

Jean P. Gaffney ◽

Inna Solomonov ◽

Maxim Levin ◽

Mordehay Klepfish ◽

...

Keyword(s):

Monoclonal Antibody ◽

Active Site ◽

Drug Targets ◽

Fas Ligand ◽

Specific Activity ◽

Matrix Metalloproteases ◽

Ductal Adenocarcinoma ◽

Enzyme Active Site ◽

Induced Apoptosis

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity.

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On the identity of vitronectin and S-protein: immunological crossreactivity and functional studies

Blood ◽

10.1182/blood.v68.3.737.bloodjournal683737 ◽

1986 ◽

Vol 68 (3) ◽

pp. 737-742

Author(s):

BR Tomasini ◽

DF Mosher

Keyword(s):

Monoclonal Antibody ◽

Serum Proteins ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Gel Filtration ◽

Membrane Attack Complex ◽

Biochemical Properties ◽

Complex Data ◽

Heparin Binding ◽

S Protein

Vitronectin (serum spreading factor), a major serum cell adhesion molecule, was compared with S-protein, the inhibitor of the C5–9 membrane attack complex. Data from the literature indicate that S- protein and vitronectin are alpha globulins with the same aminoterminal residues, amino acid compositions, and concentrations in normal plasma (150 to 250 micrograms/mL). Both proteins have been reported to interact with the thrombin-antithrombin complex. The cDNA sequences of vitronectin and S-protein were recently determined and found to be almost identical. In the present studies, rabbit-anti-S-protein and a monoclonal antibody to vitronectin both recognized 65,000- and 75,000- molecular weight (mol wt) polypeptides when plasma or serum proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose paper. The 65,000 and 75,000-mol wt polypeptides bound more avidly from serum than plasma to monoclonal anti-vitronectin or heparin coupled to agarose. The presence or absence of the polypeptides constituted a major difference between the heparin-binding proteins of serum and plasma. When complement- activated serum and unactivated serum were separated by gel filtration, vitronectin coeluted with C9 in high-mol-wt fractions of activated serum but not unactivated serum. Purified S-protein was recognized by the monoclonal antibody to vitronectin and promoted spreading of human skin fibroblasts. Both vitronectin and S-protein were degraded by thrombin. On the basis of immunological and functional, as well as biochemical, properties, therefore, S-protein and vitronectin are the same.

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THE SERO-CONVERSION AND EVALUATION OF RENAL ALTERATIONS IN DOGS INFECTED BY Leishmania (Infantum) chagasi

Revista do Instituto de Medicina Tropical de São Paulo ◽

10.1590/s0036-46652013000200007 ◽

2013 ◽

Vol 55 (2) ◽

pp. 105-112 ◽

Cited By ~ 2

Author(s):

Georgia Brenda Barros Alves ◽

Lucilene dos Santos Silva ◽

Joilson Ferreira Batista ◽

Ângela Piauilino Campos ◽

Maria das Graças Prianti ◽

...

Keyword(s):

Visceral Leishmaniasis ◽

Post Infection ◽

Serum Proteins ◽

Clinical Symptoms ◽

Periodic Acid ◽

Biochemical Tests ◽

Serological Tests ◽

Periodic Acid Schiff ◽

Infection Period ◽

Conversion Period

This study investigated the sero-conversion period in which dogs from endemic areas test positive for visceral leishmaniasis (VL) as well as the early post-infection period in which renal alterations are observed. Dogs that were initially negative for Canine Visceral Leishmaniasis (CVL) were clinically evaluated every three months by serological, parasitological and biochemical tests until sero-conversion was confirmed, and six months later a subsequent evaluation was performed. Samples of kidney tissues were processed and stained with Hematoxylin and Eosin (H&E), Periodic Acid Schiff (PAS) and Masson’s trichrome stain and lesions were classified based on the WHO criteria. Of the 40 dogs that initially tested negative for VL, 25 (62.5%) exhibited positive serological tests during the study period. Of these 25 dogs, 15 (60%) tested positive within three months, five (20%) tested positive within six months and five (20%) tested positive within nine months. The dogs exhibited antibody titers between 1:40 and 1:80 and 72% of the dogs exhibited clinical symptoms. The Leishmania antigen was present in the kidneys of recently infected dogs. We found higher levels of total protein and globulin as well as lower levels of albumin in the infected dogs when compared to the control dogs. Additionally, infected dogs presented levels of urea and creatinine that were higher than those of the uninfected dogs. Glomerulonephritis was detected in some of the dogs examined in this study. These data suggest that in Teresina, the sero-conversion for VL occurs quickly and showed that the infected dogs presented abnormal serum proteins, as well as structural and functional alterations in the kidneys during the early post-infection period.

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Blood kinetics, tissue distribution, and radioimaging of anti-common chronic lymphatic leukemia antigen (cCLLa) monoclonal antibody CLL2 in mice transplanted with cCLLa-bearing human leukemia cells

Blood ◽

10.1182/blood.v75.9.1853.1853 ◽

1990 ◽

Vol 75 (9) ◽

pp. 1853-1861 ◽

Cited By ~ 2

Author(s):

GB Faguet ◽

JF Agee ◽

JT DiPiro

Keyword(s):

Monoclonal Antibody ◽

Specific Activity ◽

Human Leukemia ◽

Chronic Lymphatic Leukemia ◽

Prolymphocytic Leukemia ◽

Normal Tissues ◽

Lymphatic Leukemia ◽

Human Leukemia Cells ◽

Glycoprotein Antigen ◽

Leukemia Antigen

Abstract The blood kinetics and biodistribution of anti-common chronic lymphatic leukemia antigen (cCLLa) monoclonal antibody (MoAb) CLL2 were assessed in mice bearing cCLLa+ tumors. The cCLLa is a 69-Kd glycoprotein antigen expressed selectively by malignant B cells in human CLL, hairy cell leukemia (HCL), and prolymphocytic leukemia. Immunoreactive 125I- CLL2 (5 micrograms/mouse, specific activity 4.3 microCi/micrograms) was injected intravenously in mice bearing HCL-derived EH xenografts, and blood kinetics and biodistribution were ascertained up to 16 days postinjection. Radioimages were also obtained up to 72 hours after injecting 10 micrograms/mouse (specific activity 50.1 microCi/micrograms) of 125I-CLL2. Distinct 125I-CLL2 blood kinetics were observed in EH engrafted compared with tumor-free mice including: a longer 125I-CLL2 T 1/2 (153 hours v 72 hours), and a considerably greater blood clearance (173 mg/h v 54.7 mg/h) with biexponential rather than monoexponential configuration; and a greater volume of antibody distribution (31,483 mg v 5,729 mg). These data suggest more rapid tissue uptake by grafted tumours. Preferential 125I-CLL2 uptake by EH tumours relative to normal tissues was observed beginning 24 hours postinjection (mean ratio, 4.2) with average peak tumor 125I-CLL2 levels of 428.7 pg/mg. 125I-CLL2 uptake selectivity by EH tumor cells was also supported by: (1) negligible 125I-CLL2 uptake by cCLLa- Molt-4 xenografts (average 29.1 pg/mg 24 hours postinjection); (2) background uptake of cCLLa-irrelevant MoAb 131I-LEU1 by CD5- EH xenografts (average 31.4 pg/mg 48 hours postinjection); and (3) by scintigraphy. The EH xenograft mouse model might be useful to ascertain preclinically the anti-tumor effect of anti-cCLLa MoAbs and of their conjugated derivatives.

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Rat hepatic bile acid sulfotransferase: enzyme response to androgens and estrogens

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1987.252.2.g276 ◽

1987 ◽

Vol 252 (2) ◽

pp. G276-G280

Author(s):

R. H. Collins ◽

L. Lack ◽

P. G. Killenberg

Keyword(s):

Monoclonal Antibody ◽

Bile Acid ◽

High Performance ◽

Protein Concentration ◽

Specific Activity ◽

Antibody Activity ◽

Hepatic Bile ◽

Reactive Protein ◽

Effect Of Treatment ◽

Relative Molecular Weight

Rat liver bile sulfotransferase activity can be divided into a fraction that reacts with a monoclonal antibody (PK1B) and another fraction that does not. This work was performed to analyze the known response of hepatic bile acid sulfotransferase activity to androgens and estrogens by determining the effect of treatment on the proportion of bile acid sulfotransferase activity that possessed the epitope for PK1B monoclonal antibody. Activity in treated animals was further characterized by high-performance liquid chromatography (HPLC) analysis following purification by PK1B-immunoadsorption chromatography. The results indicate that estrogens and androgens affect the subset of enzyme activity that has the PK1B epitope more than the population that does not. HPLC demonstrates that increases and decreases in activity that follow treatment with androgens and estrogens are mirrored by the proportion of the PK1B-reactive protein that exhibits a relative molecular weight (Mr) greater than 170,000. Radial immunodiffusion assays of hepatic supernatant using a polyclonal antibody raised against PK1B-reactive bile acid sulfotransferase show that changes in specific activity that follow treatment are the result of changes in enzyme protein concentration.

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Destruction of Immature Erythrocytes Measured by Bilirubin Excretion

Blood ◽

10.1182/blood.v33.5.717.717 ◽

1969 ◽

Vol 33 (5) ◽

pp. 717-726

Author(s):

KIYOYASU NAGAI ◽

EIZO KAKISHITA

Keyword(s):

Normal State ◽

General Circulation ◽

Direct Evidence ◽

Specific Activity ◽

Total Radioactivity ◽

Sprague Dawley ◽

Wistar Strain ◽

Bilirubin Excretion ◽

Sprague Dawley Rats ◽

Bile Fistula

Abstract The change in radioactivity of bilirubin with time was measured after injection of glycine 2-14C into rats with a bile fistula. The total radioactivity and specific activity of bilirubin were abnormally high in rats of which erythropoiesis was increased following hemorrhage. When the newly formed erythrocytes labeled with glycine 2-14C in the peak of reticulocytosis were transfused into rats with a bile fistula, the excretion of labeled bilirubin increased rapidly. The amount of newly formed erythrocytes destroyed within 7 days after transfusion into normal animals was calculated as 4.4 per cent of the total erythrocytes formed during maximum reticulocytosis and 1.1 per cent of those formed in the normal state in Sprague Dawley rats, while in the Wistar strain, the values were 13.4 per cent and 2.2 per cent, respectively. These results provide direct evidence for increased production of short-lived erythrocytes during enhanced erythropoiesis, and the hemolysis of newly formed erythrocytes soon after they reach the general circulation may contribute to the production of shunt bilirubin.

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Purification, characterization and inhibition of human skin collagenase

Biochemical Journal ◽

10.1042/bj1690265 ◽

1978 ◽

Vol 169 (2) ◽

pp. 265-276 ◽

Cited By ~ 89

Author(s):

David E. Woolley ◽

Robert W. Glanville ◽

Dennis R. Roberts ◽

John M. Evanson

Keyword(s):

Human Skin ◽

Culture Medium ◽

Serum Proteins ◽

Specific Activity ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Gel Filtration ◽

Polyacrylamide Gel ◽

Collagen Fibrils ◽

Ph Optimum

1. The neutral collagenase released into the culture medium by explants of human skin tissue was purified by ultrafiltration and column chromatography. The final enzyme preparation had a specific activity against thermally reconstituted collagen fibrils of 32μg of collagen degraded/min per mg of enzyme protein, representing a 266-fold increase over that of the culture medium. Electrophoresis in polyacrylamide disc gels showed it to migrate as a single protein band from which enzyme activity could be eluted. Chromatographic and polyacrylamide-gel-elution experiments provided no evidence for the existence of more than one active collagenase. 2. The molecular weight of the enzyme estimated from gel filtration and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis was approx. 60000. The purified collagenase, having a pH optimum of 7.5–8.5, did not hydrolyse the synthetic collagen peptide 4-phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-d-Arg-OH and had no non-specific proteinase activity when examined against non-collagenous proteins. 3. It attacked undenatured collagen in solution at 25°C, producing the two characteristic products TCA(¾) and TCB(¼). Collagen types I, II and III were all cleaved in a similar manner by the enzyme at 25°C, but under similar conditions basement-membrane collagen appeared not to be susceptible to collagenase attack. At 37°C the enzyme attacked gelatin, producing initially three-quarter and one-quarter fragments of the α-chains, which were degraded further at a lower rate. As judged by the release of soluble hydroxyproline peptides and electron microscopy, the purified enzyme degraded insoluble collagen derived from human skin at 37°C, but at a rate much lower than that for reconstituted collagen fibrils. 4. Inhibition of the skin collagenase was obtained with EDTA, 1,10-phenanthroline, cysteine, dithiothreitol and sodium aurothiomaleate. Cartilage proteoglycans did not inhibit the enzyme. The serum proteins α2-macroglobulin and β1-anti-collagenase both inhibited the enzyme, but α1-anti-trypsin did not. 5. The physicochemical and enzymic properties of the skin enzyme are discussed in relation to those of other human collagenases.

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SELECTIVE RELEASE OF CONTENT FROM MICROSOMAL VESICLES WITHOUT MEMBRANE DISASSEMBLY

The Journal of Cell Biology ◽

10.1083/jcb.61.3.789 ◽

1974 ◽

Vol 61 (3) ◽

pp. 789-807 ◽

Cited By ~ 49

Author(s):

Gert Kreibich ◽

David D. Sabatini

Keyword(s):

Serum Proteins ◽

Specific Activity ◽

Membrane Vesicles ◽

Rat Serum ◽

Coomassie Blue ◽

Specific Subset ◽

Low Levels ◽

Almost All

Rough and smooth microsomes were shown to have similar sets of polypeptide chains except for the proteins of ribosomes bound to the rough endoplasmic reticulum (ER). More than 50 species of polypeptides were detected by acrylamide gel electrophoresis, ranging in molecular weight from 10,000 to approximately 200,000 daltons. The content of rough and smooth microsomes was separated from the membrane vesicles using sublytic concentrations of detergents and differential centrifugation. A specific subset of proteins which consisted of approximately 25 polypeptides was characteristic of the microsomal content. Some of these proteins showed high rates of in vivo incorporation of radioactive leucine or glucosamine, but several others incorporated only low levels of radioactivity within short labeling intervals and appeared to be long-term residents of the lumen of the ER. Seven polypeptides in the content subfractions, including serum albumin, contained almost 50% of the leucine radioactivity incorporated during 5 min and cross-reacted with antiserum against rat serum. Almost all microsomal glycoproteins were at least partly released with the microsomal content. Smooth microsomes contained higher levels of albumin than rough microsomes, but after short times of labeling with [3H]leucine the specific activity of albumin in the latter was higher, supporting the notion that newly synthesized serum proteins are transferred from rough to smooth portions of the ER. On the other hand, after labeling for 30 min with [3H]glucosamine, smooth microsomes contained higher levels of radioactivity than rough microsomes. This would be expected if glycosidation of newly synthesized polypeptides proceeds during their transit through ER cisternae. The labeling pattern of membrane proteins in microsomes obtained from animals which received three daily injections of [3H]leucine, the last administered 1 day before sacrifice, followed the intensity of bands stained with Coomassie blue, with a main radioactive peak corresponding to cytochrome P 450. After the long-term labeling procedure most content proteins had low levels of radioactivity; this was especially true of serum proteins which were highly labeled after 30 min.

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Phase I/II study of iodine 131-labeled monoclonal antibody A33 in patients with advanced colon cancer.

Journal of Clinical Oncology ◽

10.1200/jco.1994.12.8.1561 ◽

1994 ◽

Vol 12 (8) ◽

pp. 1561-1571 ◽

Cited By ~ 88

Author(s):

S Welt ◽

C R Divgi ◽

N Kemeny ◽

R D Finn ◽

A M Scott ◽

...

Keyword(s):

Monoclonal Antibody ◽

Colon Cancer ◽

Antitumor Activity ◽

Phase I ◽

Specific Activity ◽

Gastrointestinal Symptoms ◽

Whole Body ◽

Heavily Pretreated ◽

Pretreated Patients ◽

Iodine 131

PURPOSE A phase I/II study was designed to determine the maximum-tolerated dose (MTD) of iodine 131-labeled monoclonal antibody (mAb) A33 (131I-mAb A33) administered intravenously, its limiting organ toxicity, and its radioisotope retention in tumors, and to develop preliminary evidence of antitumor activity. PATIENTS AND METHODS Patients (N = 23) with colorectal cancer who had failed to respond to conventional chemotherapy but had not received prior radiotherapy were treated with escalating doses of 131I-mAb A33. Three or more patients were entered at each dose level, starting at 30 mCi/m2, with increments of 15 mCi/m2 to a maximal dose of 90 mCi/m2. Radiolabeling was performed to maintain a specific activity of 30 mCi/m2/4 mg mAb A33 (projected maximum, 15 mCi/mg). Patients were under strict isolation precautions until whole-body radiation levels decreased to less than 5 mrem/h at 1 m. Serial radioimmunoscintigrams were performed in some cases for up to 3 weeks after 131I-mAb A33 administration. RESULTS All 20 patients with radiologic evidence of disease showed localization of radioisotope to sites of disease. Two patients with elevated carcinoembryonic antigen (CEA) levels and negative radiologic tests did not have positive antibody scans. One patient with a small-bowel cancer also had a negative antibody scan. The major toxicity was hematologic and was more pronounced in patients with compromised bone marrow due to prior chemotherapy. Of five patients who received 78 to 84 mCi/m2 131I-mAb A33, one had grade 3 and one grade 4 toxicity; of six patients treated with 86 to 94 mCi/m2 131I-mAb A33, two had grade 4 and one grade 1 toxicity. The MTD was determined to be 75 mCi/m2 in these heavily pretreated patients. Although the isotope showed variable uptake in the normal bowel, gastrointestinal symptoms were mild (n = 8) or absent. No major responses were observed; however, three patients had evidence of mixed responses, and CEA levels decreased in two patients without clinical or radiologic measurable disease. Immunoreactivity of radiolabeled mAb A33 decreased at the highest dose levels in preparations in which specific activity exceeded 18 mCi/mg. CONCLUSION The A33 antigen appears to be a promising target for radioimmunotherapy of colon cancer. The modest antitumor activity of 131I-mAb A33 in heavily pretreated patients is encouraging because of its lack of toxicity in the bowel, the only antigen-positive normal tissue.

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Isolation and Characterization of a Novel Bacterium from the Marine Environment for Trichloroacetic Acid Bioremediation

Applied Sciences ◽

10.3390/app10134593 ◽

2020 ◽

Vol 10 (13) ◽

pp. 4593 ◽

Cited By ~ 2

Author(s):

Mahshid Heidarrezaei ◽

Hoofar Shokravi ◽

Fahrul Huyop ◽

Seyed Saeid Rahimian Koloor ◽

Michal Petrů

Keyword(s):

16S Rrna ◽

Trichloroacetic Acid ◽

Specific Activity ◽

Morphological Observation ◽

Rrna Gene ◽

Industrial Chemicals ◽

Halogenated Compounds ◽

Biochemical Tests ◽

Isolation And Characterization ◽

Halogenated Compound

Halogenated compounds are an important class of environmental pollutants that are widely used in industrial chemicals such as solvents, herbicides, and pesticides. Many studies have been carried out to explore the biodegradation of these chemicals. Trichloroacetic acid (TCA) is one of the main halogenated compounds that are carcinogenic to humans and animals. The bacterium was isolated from the northern coastline of Johor Strait. In this study, the ability of strain MH2 to biodegrade TCA was evaluated by a growth experiment and dehalogenase enzyme assay. The growth profile of the isolated strain was examined. The doubling time for L. boronitolerans MH2 was found to be 32 h. The release of chloride ion in the degradation process was measured at 0.33 × 10−3 ± 0.03 mol∙L−1 after 96 h when the growth curve had reached its maximum within the late bacterial exponential phase. The results showed that the strain had a promising ability to degrade TCA by producing dehalogenase enzyme when cell-free extracts were prepared from growth on TCA as the sole carbon source with enzyme-specific activity, 1.1 ± 0.05 µmolCl−min−1∙mg−1 protein. Furthermore, the morphological, and biochemical aspects of the isolated bacterium were studied to identify and characterize the strain. The morphological observation of the isolated bacterium was seen to be a rod-shaped, Gram-positive, motile, heterotrophic, and spore-forming bacterium. The amplification of the 16S rRNA and gene analysis results indicated that the isolated bacterium had 98% similarity to Lysinibacillus boronitolerans. The morphological and biochemical tests supported the 16S rRNA gene amplification. To the best of the authors’ knowledge, this is the first reported case of this genus of bacteria to degrade this type of halogenated compound.

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