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Biomedical subjects

J R Murphy

Publications and source records attributed to J R Murphy.

At least 199 records · Page 11Linked to original sources
PubMed1986

Genetic construction, expression, and melanoma-selective cytotoxicity of a diphtheria toxin-related alpha-melanocyte-stimulating hormone fusion protein.

The structural gene for diphtheria toxin, tox, has been modified at its Sph I site by the introduction of an oligonucleotide linker encoding a unique Pst I restriction endonuclease site and a synthetic oligonucleotide encoding alpha-melanocyte-stimulating hormone (alpha-MSH). The resulting fusion gene directs the expression of a diphtheria toxin-related alpha-MSH hybrid protein in which the diphtheria toxin receptor-binding domain has been replaced with alpha-MSH sequences. The chimeric toxin has been partially purified from periplasmic extracts of recombinant Escherichia coli K-12 and has been found to be selectively toxic for alpha-MSH receptor-positive human malignant melanoma NEL-M1 cells in vitro.

ADP Ribose Transferases↗
PubMed1986

Comparison of alternating to nonalternating chemotherapy regimens for non-small cell lung cancer.

Forty-seven patients with stage III lung cancer from four institutions in the Denver area were entered in a study comparing two regimens of chemotherapy. The patients were randomized into two groups: Group A received lomustine, cyclophosphamide, vincristine, cisplatin, and doxorubicin monthly; Group B received the five-drug regimen on Months 1, 3, and 5 and received 5-FU by constant infusion, methotrexate, and mitomycin on Months 2, 4, and 6. The age, extent of disease, Karnofsky score, prior therapy, and average number of chemotherapy courses received in each group were comparable. The median survival in Group A was 265 days and in Group B was 163 days (P greater than 0.25). There does not seem to be an advantage in survival in patients who are treated with the eight-drug regimen over the five-drug regimen.

Adenocarcinoma↗
PubMed1986

On multiple comparisons in the randomization analysis of growth and response curves.

Two multiple-comparisons procedures are suggested for supplementing randomization analysis of growth and response curves. One controls the experimentwise Type I error rate for all possible contrast curves via an extension of the Scheffé method. The other controls a family of Type I error rates via a stepwise testing procedure. Both can be approximated by standard F tests without costly recomputation of all of the test statistics for a large number of permutations.

Biometry↗
PubMed1985

Pre-clinical toxicology of nitazoxanide--a new antiparasitic compound.

The acute and subchronic toxicological effects of nitazoxanide were investigated at levels near and in excess of the therapeutic dose in rats, mice, dogs and cats. Single oral gavage doses of 0.625-10 g per kg body weight were administered to rats and mice. Single oral doses of 1-10 g per kg body weight were administered in capsules to dogs and cats. Acute oral LD50 values were greater than 10 g kg-1 in rats, dogs and cats, and 1.4 g kg-1 in mice. Systemic toxicity was evaluated in a repeated dose study in rats at doses of 50, 150 and 450 mg per kg per day for 14 weeks. The highest dose group exhibited intense salivation, increased liver and spleen weight, and decreased thymus weights. Variances between control and treated organ weights were not confirmed by histopathological evaluation. Nitazoxanide was negative when tested in the Ames Salmonella assay using five tester strains with and without metabolic activation at levels from 1 to 100 mg per plate. The drug was also shown to be non-irritating in a test for eye irritation potential.

Animals↗
PubMed1985

Cloning and characterization of the hemolysin determinants from Vibrio cholerae RV79(Hly+), RV79(Hly-), and 569B.

The Hly region from the chromosome of Vibrio cholerae El Tor strain RV79(Hly-) and the nonhemolytic classical strain 569B were cloned into plasmid vector pBR322. Escherichia coli K-12 transformants possessing these recombinant plasmids were nonhemolytic and were detected with a 32P-labeled hly-specific DNA probe. Restriction endonuclease Sau3AI digestions of the cloned hly loci of two independently obtained RV79(Hly+) convertants, when compared with the digests of cloned RV79(Hly-) loci, revealed that an apparent alteration (10 to 15 base pairs) had occurred. In contrast, an apparent 20-base-pair deletion was present in the cloned hly locus of the classical biotype V. cholerae strain 569B. Maxicell analysis and immunoprecipitation of labeled proteins of E. coli which are encoded by the cloned hly loci of RV79(Hly+) and from nuclease BAL 31-deleted plasmids, as well as immunoprecipitation of [35S]methionine-labeled V. cholerae proteins, suggest that the hemolysin is an 84,000-dalton polypeptide.

Base Sequence↗
PubMed1985

Characterization of the diphtheria tox transcript in Corynebacterium diphtheriae and Escherichia coli.

Transcription of the tox gene in lysogenic Corynebacterium diphtheriae strains C7(beta tox+), C7 (gamma tox) and the hypertoxigenic PW8 (omega tox+) was analyzed and compared with transcription of the C. diphtheriae tox gene in the recombinant strain Escherichia coli (pDT201). In all cases S1 nuclease mapping localized the 5' terminus of the tox mRNA to a site 8 or 9 base pairs (bp) downstream of a region similar to the -10 consensus sequence of E. coli promoters. In C. diphtheriae the tox transcript was observed only in strains that were grown under iron-limiting conditions; in the presence of excess iron, transcription beyond bp 38 of the tox coding region was not observed. In contrast, in E. coli(pDT201) tox was expressed at equivalent levels in both iron-depleted and iron-supplemented media. The DNA insertion in the tox gene of the nontoxigenic corynephage gamma was found to occur at bp 54 of the tox coding region. The insertion event resulted in the duplication of a 7-bp target sequence, and the ends of the insert were found to constitute an imperfect inverted repeat of approximately 26 bp. Transcription from the tox promoter in C7(gamma tox) was found to initiate at the same nucleotides as in C7(beta tox+), PW8, and E. coli(pDT201) and remained sensitive to iron inhibition. These observations are discussed in relation to the mechanism of iron-mediated regulation of the tox gene.

Base Sequence↗
PubMed1985

Detection and physical map of a omega tox+-related defective prophage in Corynebacterium diphtheriae Belfanti 1030(-)tox-.

A library of chromosomal DNA from Corynebacterium diphtheriae Belfanti 1030(-)tox- was cloned in the lambda phage vector EMBL4 and screened for sequences homologous to corynephage omega tox+ and the attB1-attB2 region of the C7(-)tox- chromosome. Two portions of the 1030(-)tox- chromosome, 35 and 30.5 kilobases long which contain, respectively, the entire region homologous to corynephage omega tox+ and the attB1-attB2 sites, were mapped with the restriction endonucleases BamHI and EcoRI. Chromosomal DNA from 1030(-)tox- was shown to contain a 15.5-kilobase region that was homologous to ca. 42% of the corynephage omega tox+ genome. These sequences were found to hybridize to three regions of the phage genome and do not contain either the diphtheria tox operon or the attP site. These sequences are distant from the chromosomal region that contains the attB1-attB2 sites. Moreover, unlike other known defective prophages, the physical map of this prophage starts at the cos site and is colinear with the vegetative phage map. The 30.5-kilobase region of the 1030(-)tox- chromosome, which contains the attB1-attB2 sites, has a central core region that is almost identical to the corresponding region of the C7(-)tox- chromosome; however, the flanking sequences in these two strains of C. diphtheriae are different.

Bacteriophages↗
PubMed1985

Malignant placental site trophoblastic tumor.

Placental site trophoblastic tumor is a rare variant of trophoblastic disease. The malignant form of this disease with metastasis and death is even more infrequent, as is evident from only five cases in the literature. A 30-year-old black woman who died from this disease is presented. Patients with metastases are at extremely high risk with no reported survivors and must be treated aggressively with chemotherapy and cytoreductive surgery. This type of trophoblastic tumor apparently exhibits a different biologic behavior as compared with choriocarcinoma.

Adult↗
PubMed1984

Monoclonal antibody analysis of diphtheria toxin--I. Localization of epitopes and neutralization of cytotoxicity.

Forty-three hybridoma cell lines producing monoclonal antibody to diphtheria toxin were isolated. Based upon their reactivity with various fragments of the toxin and mutant toxin-related proteins, the monoclonal antibodies were subdivided into 10 groups which recognize at least 10 distinct epitopes on the toxin molecule. Specific antibodies directed against both fragments A and B were found to neutralize cytotoxicity, both in vitro and in vivo. Neutralization was found to correlate with antibody-mediated inhibition of toxin binding to its eukaryotic cell surface receptor. The results presented suggest that the Pro378 and/or Gly431 of mature toxin are part of, or close to, the toxin receptor-binding domain. In addition, antigenic determinants in the C-terminal portion of fragment A, as well as a portion of the toxin defined by the tox-3 and tox-45 nonsense mutations (i.e. ca 31,000-42,000 daltons) appear to be juxtaposed to the receptor-binding domain and may form a secondary binding region.

Animals↗
PubMed1984

Monoclonal antibody analysis of diphtheria toxin--II. Inhibition of ADP-ribosyl-transferase activity.

Monoclonal antibodies directed against the enzymatically active A-fragment of diphtheria toxin were used to investigate further the structure-function relationships within fragment A. Of 16 such antibodies, all but two were directed against epitopes located within the carboxy-terminal 30-40 amino acids of fragment A. Interestingly, the antibodies recognize several epitopes in this small region and varied considerably in their effects on toxin functions. With regard to their effects on the enzymatic activity of fragment A, three types of antibodies were found: (1) antibodies which bind fragment A but fail to inhibit its ADP-ribosyltransferase activity, (2) antibodies which completely inhibit enzyme activity, and (3) antibodies which interact with fragment A to yield antigen-antibody complexes of diminished activity. The results are consistent with location of the catalytic center of fragment A within its carboxy-terminal ca 4000 dalton region.

Animals↗
PubMed1984

Characteristics of Rickettsia mooseri infection of normal and immune mice.

Rickettsia mooseri infection initiated by subcutaneous injection has been studied in BALB/c mice with the objective of developing a model for the study of immune mechanisms. Characterization of infection included the following: measurement of the replication, dissemination, and clearance of rickettsiae; measurement of correlates of the immune response, including humoral antibody, hypersensitivity to subcutaneously inoculated rickettsial antigen, and activation of nonspecific macrophage microbicidal capacity; and measurement of resistance to a second homologous challenge. Local infection at the site of subcutaneous injection progressed through day 5 and was controlled by day 7. Systemic infection as determined by the presence of rickettsiae in spleen was first detected on day 7 and progressed through day 14; however, rickettsiae persisted in this organ at reduced numbers through at least day 28. Control of the local infection at the site of subcutaneous injection occurred at about the time humoral antibodies and hypersensitivity reactions to subcutaneously injected rickettsial antigens became demonstrable and was paralleled by a capacity to resist homologous subcutaneous challenge at a site distant from that of the primary infection. Systemic infection progressed in spite of this acquired immune capacity and was controlled in the spleen in parallel with the development of enhanced macrophage microbicidal capacity in the liver. The results show that an acquired immunity is capable of restricting rickettsial growth at subcutaneous sites at a time when rickettsiae are increasing in titer in deep organs.

Animals↗
PubMed1984

Characteristics of lymphoid cells that adoptively transfer immunity to Rickettsia mooseri infection in mice.

The capacity of adoptively transferred immune lymphoid cells or passively transferred immune serum to alter the course of an established Rickettsia mooseri (R. typhi) infection in the spleen was evaluated in BALB/c mice. Immune cells, but not immune serum, controlled the established infection. An effective lymphocyte was a T-cell which had to possess a capacity to divide.

Animals↗
PubMed1984

Molecular cloning of the hemolysin determinant from Vibrio cholerae El Tor.

Vibrio cholerae El Tor RV79 is phenotypically nonhemolytic; however, strongly hemolytic convertants are occasionally observed on blood agar plates. We have cloned DNA sequences corresponding to the hemolysin determinant from RV79 (Hly+) in the lambda L47.1 and pBR322 vectors. A 2.3-kilobase fragment of V. cholerae DNA was found to be necessary for hemolytic activity. This cloned DNA sequence was used as a probe in Southern blot hybridization analysis of chromosomal restriction digests of a variety of El Tor and classical biotype V. cholerae strains. In all cases, DNA fragments with the same electrophoretic mobilities hybridized to the Hly probe. The results presented demonstrate that the cloned hemolysin determinant is the hly locus. By using mutator vibriophage VcA-3 insertion to promote high-frequency transfer, the hly locus was mapped between arg and ilv on the V. cholerae RV79 chromosome.

Base Sequence↗
PubMed1984

Host defenses in murine malaria: humoral immunity to Plasmodium berghei in mice.

Humoral immunity to Plasmodium berghei infection of F1 hybrid B6D2 (C57B1/6 X DBA/2) mice was investigated using an immune serum prepared from mice which survived a lethal challenge of erythrocytic stage P. berghei because of previous vaccination with formalin-killed P. berghei. Immune serum, but not normal serum, if injected intraperitoneally or intravenously soon after infection, during rapidly increasing parasitemia, transiently inhibited the progress of infection in a pattern which was directly related to the dose and timing of serum injection. However, the level of restriction of parasitemia caused by the intravenous injection of 1 ml of immune serum could not be improved upon by subsequent additional 1-ml injections of the same serum, and parasitemia in all mice, whether delayed by immune serum or not, ultimately progressed to death. The protective capacity of the immune serum was markedly specific for plasmodial species but absorption with formalin-killed homologous parasites failed to remove the protective component. No variants of P. berghei insensitive to inhibition by immune serum were detected. The results support the view that humoral immunity to P. berghei may be mediated by more than one soluble factor and effected through at least two different pathways.

Animals↗
PubMed1983

Cloned diphtheria toxin fragment A is expressed from the tox promoter and exported to the periplasm by the SecA apparatus of Escherichia coli K12.

The diphtheria toxin fragment A-related polypeptide encoded by the recombinant plasmid pDT201 (Leong, D., Coleman, K. D., and Murphy, J. R. (1983) Science 220, 515-517) and expressed in Escherichia coli K12 was found to have an electrophoretic mobility in sodium dodecyl sulfate-polyacrylamide gels corresponding to 28,500 daltons. Immunoprecipitation experiments revealed that the mature form of the fragment A-related polypeptide was exported to the periplasmic compartment of E. coli K12 harboring plasmid pDT201. The polypeptide was sensitive to trypsin "nicking." Following treatment an Mr = 24,000 fragment was released which co-electrophoresed with fragment A of diphtheria toxin. The precursor form (Mr = 31,000) of the fragment A-related polypeptide was found to accumulate in the cytoplasmic fraction of the temperature-sensitive secretion-defective strain of E. coli K12 under nonpermissive conditions. We further demonstrate that a 263-base pair HaeIII fragment of pDT201 which carries the putative diphtheria tox promoter has promoter activity in E. coli K12.

Autoradiography↗