HPLC in nucleic acids research.
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Chromatography, High Pressure Liquid↗
Biomedical subjects
J A Thompson
Publications and source records attributed to J A Thompson.
Chromosomal localization of the carcinoembryonic antigen gene family and differential expression in various tumors.
Carcinoembryonic antigen (CEA) is a glycoprotein which is important as a tumor marker for a number of human cancers. It is a member of a gene family comprising about 10 closely related genes. In order to characterize mRNAs transcribed from individual genes we have identified by DNA and RNA hybridization experiments, gene-specific sequences from the 3' noncoding regions of CEA, and of nonspecific cross-reacting antigen (NCA) mRNAs, which have been recently cloned. With these probes, CEA mRNAs with lengths of 3.5 and 3.0 kilobases and an NCA mRNA species of 2.5 kilobases were identified in various human tumors. A 2.2-kilobase mRNA species, however, could only be detected in leukocytes of patients with chronic myeloid leukemia by hybridization with a probe from the immunoglobulin-like repeat domain of CEA. This region is known to be very similar among the various members of the CEA gene family, and indeed the probe hybridizes with all four mRNA species. In situ hybridization with a cross-hybridizing probe from the NCA gene localized the members of the CEA gene family to the short and to the long arm of chromosome 19. In addition, a CEA cDNA probe was found to hybridize to the long arm of chromosome 19 only.
Antigens, Neoplasm↗
4-Hydroxybutyric aciduria in a patient without ataxia or convulsions.
A child presenting with mild psychomotor retardation, hypotonia, microcephaly and hyperkinesis is described. Urinary organic acid analysis by combined gas chromatography-mass spectrometry revealed 4-hydroxybutyric aciduria. Succinic semialdehyde dehydrogenase activity in extracts of white cells derived from the patient was less than 10% of control values.
Ataxia↗
DNA affinity labeling of adenovirus type 2 upstream promoter sequence-binding factors identifies two distinct proteins.
A rapid affinity labeling procedure with enhanced specificity was developed to identify DNA-binding proteins. 32P was first introduced at unique phosphodiester bonds within the DNA recognition sequence. UV light-dependent cross-linking of pyrimidines to amino acid residues in direct contact at the binding site, followed by micrococcal nuclease digestion, resulted in the transfer of 32P to only those specific protein(s) which recognized the binding sequence. This method was applied to the detection and characterization of proteins that bound to the upstream promoter sequence (-50 to -66) of the human adenovirus type 2 major late promoter. We detected two distinct proteins with molecular weights of 45,000 and 116,000 that interacted with this promoter element. The two proteins differed significantly in their chromatographic and cross-linking behaviors.
Adenoviruses, Human↗
Influence of dose and duration of infusion of interleukin-2 on toxicity and immunomodulation.
The purpose of this study was to investigate the effect of dose and duration of infusion of recombinant interleukin-2 (IL-2) on toxicity and immunomodulation. In a phase I/II study, IL-2 was administered intravenously (IV) daily for five consecutive days every other week for 4 weeks of treatment to 23 patients with progressive melanoma, renal, colon, or ovarian cancer by one of four regimens: groups I and II received 3 X 10(5) U/m2/d by two-hour or 24-hour infusion, respectively; groups III and IV received 3 X 10(6) U/m2/d by two-hour or 24-hour infusion, respectively. In a subsequent study, six patients (group V) received a single priming cycle of daily IL-2 for five days at 3 X 10(6) U/m2/d in divided 15-minute infusions every eight hours, before undergoing leukapheresis for lymphokine-activated killer (LAK) cell generation. Toxicity was mild with 3 X 10(5) U/m2/d, but severe chills and fever, moderate hypotension (not requiring IV pressors), and weight gain were observed with 3 X 10(6) U/m2/d. Toxicity was also related to the duration of infusion. In group IV (continuous infusion), fluid retention, weight gain, and azotemia were more frequent and severe than in groups III or V, in which the same total dose was administered by two-hour infusion or in three divided 15-minute infusions. IL-2 induced rebound lymphocytosis, which was directly dose-related and significantly higher in group IV (continuous infusion) than in groups III or V. Dramatic increases in the percentage and absolute number of cells expressing the IL-2 receptor were also most pronounced in group IV. With the higher dose of IL-2, LAK cells appeared in the circulation, and natural killer (NK) cytotoxicity was augmented. The results showed that the toxicity and immunomodulation by IL-2 are dose-dependent and are maximal by continuous infusion compared with two-hour or divided every eight hours infusions.
Adult↗
Applications of percutaneous transluminal coronary angioplasty in cardiac transplantation. Preliminary results in five patients.
Atherosclerotic coronary artery disease is the major cause of late cardiac transplant failure secondary to silent ischemia and infarction. To increase the longevity of cardiac homografts, we performed percutaneous transluminal coronary angioplasty (PTCA) in five male patients (aged 45 +/- 7 years, mean +/- SEM); 17 lesions were dilated during eight procedures 83 +/- 11 months after cardiac transplant. PTCA was successful (greater than or equal to 20% change in vessel diameter) in 13 of 17 (76%) lesions (the degree of prePTCA stenosis was 84% +/- 3% vs. 40% +/- 4% postPTCA; p less than or equal to 0.01). Multiple PTCA procedures were performed for progressive coronary artery disease in two patients; in one patient, two procedures were 13 months apart, and, in the second patient, another three procedures were 2 and 6 months apart. Indications for PTCA included reversible thallium perfusion defects, segmental left ventricular wall-motion abnormalities, or both in the distribution of proximal coronary artery stenoses. No deterioration occurred in the four unsuccessful PTCA attempts (two patients with initial total occlusion, and two patients in whom the lesion could not be crossed with a guidewire). Noninvasive evidence of ischemia was improved immediately after PTCA in all cases. Three patients remain alive 5, 7, and 11 months, respectively, after PTCA without evidence of new ischemia. One patient died 39 months after his first PTCA, while another patient was retransplanted 8 months after the first PTCA. Thus, PTCA can be performed in cardiac transplant patients with proximal major vessel coronary artery disease and may prolong cardiac homograft function.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
Inter- and intramolecular deuterium isotope effects on the cytochrome P-450-catalyzed oxidative dehalogenation of 1,1,2,2-tetrachloroethane.
The oxidation of 1,1,2,2-tetrachloroethane to dichloroacetic acid was investigated with rat liver microsomes and purified cytochrome P-450. Deuterium substitution had no effect on Km values, but both the inter- and intramolecular isotope effects (kH/kD) on Vmax were in the range 5.7-6.1. The equivalence of the inter- and intramolecular values indicates that 6.0 may be a good estimate of the intrinsic isotope effect. The intermolecular kH/kD value for the conversion of 1,1,2,2-trichloroethane and its 1-2H analog to chloroacetic acid was 5.5. These data, and the finding that 1 atom of 18O was incorporated into the product when TCEA was oxidized in an 18O2 atmosphere, support an oxidative dechlorination mechanism that involves hydrogen atom abstraction by the P-450 intermediate oxo complex.
Animals↗
Recombinant interleukin 2 toxicity, pharmacokinetics, and immunomodulatory effects in a phase I trial.
Twenty-two patients with refractory malignancies were treated with four escalating weekly doses of recombinant interleukin 2 (IL2), given either i.v. by 2- or 24-h infusion, or s.c. A 1-wk washout period between each dose of IL2 was provided for the evaluation for pharmacokinetic and immunomodulatory effects. The maximum i.v. dose was 30 X 10(6) units; the dose-limiting toxicities were fever, flu-like symptoms, and hypotension. The maximum s.c. dose was 3 X 10(6) because of volume limitations with s.c. injection. No tumor regression was seen. During infusions of 3 X 10(6) units over 2 h or 24 h, serum IL2 levels greater than or equal to 223 units/ml or 16 units/ml were maintained, respectively; with s.c. injection of 3 X 10(6) units, levels greater than 20 units/ml were maintained for 9 h. Marked lymphopenia was observed 24 h after the initiation of IL2 doses which was completely reversible when measured prior to the next dose. The lymphopenia was nonselective; T- and B-lymphocytes decreased in an IL2 dose-dependent manner, without consistent change in the OKT4:OKT8 ratio. No change was detected in monocyte expression of HLA-Dr or T-cell expression of the IL2 receptor. The in vitro generation of lymphokine-activated killer cytotoxicity decreased sharply and transiently shortly after i.v. doses. Mitogen responsiveness, delayed-type hypersensitivity, natural killer cytotoxicity, and mixed-lymphocyte reactivity were unchanged or decreased transiently shortly after IL2 doses. These studies help define the bioavailability of IL2 by i.v. or s.c. routes, and they will aid in the design of studies utilizing daily doses of IL2.
Antibodies↗
Antigen-driven T cell clones can proliferate in vivo, eradicate disseminated leukemia, and provide specific immunologic memory.
The aim of the current study was to determine the ability of antigen-driven cloned helper cell independent cytotoxic T lymphocytes (HITc) to proliferate and to survive in vivo and to mediate tumor therapy. The HITc clone utilized (denoted 1.B6) was specifically cytolytic to FBL-3, a syngeneic Friend virus-induced murine leukemia. Activation in vitro (48 hr) with FBL-3 induced secretion of interleukin 2 (IL 2), expression of IL 2 receptors (IL 2R), and in vitro proliferation. These cells could be "rested" for several weeks without stimulation, which resulted in reduced expression of IL 2R; however, restimulation with antigen resulted in reinduction of IL 2R and proliferation. The ability of cloned HITc to proliferate and to survive in vivo was examined in cyclophosphamide (CY) pretreated donor mice congenic for the Thy-1 gene. Adoptively transferred cloned HITc could be found in large numbers, and were widely distributed in vivo 1 wk after transfer. In tumor therapy, 1.B6 cells when injected into a site of tumor (i.p.) and used as an adjunct to CY were effective against disseminated FBL-3. In this circumstance, cloned 1.B6 cells could be recovered from cured mice 125 days after transfer and were shown to specifically lyse tumor and proliferate in vitro in response to FBL-3. Thus as an adjunct to CY, tumor-specific cloned HITc are capable of eradicating disseminated leukemia, persisting long-term in vivo, and providing specific immunologic memory.
Animals↗
Cyclosporine-induced pericardial effusion after cardiac transplantation.
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Clinical Trials as Topic↗
Resurgence of acute rheumatic fever in the intermountain area of the United States.
We describe an outbreak of acute rheumatic fever that occurred in the intermountain area centered in Salt Lake City, Utah. Seventy-four children meeting the modified Jones criteria for the diagnosis of acute rheumatic fever were evaluated by the staff at Primary Children's Medical Center, Salt Lake City, from January 1985 through June 1986. This represents an eightfold increase over the average annual incidence at this hospital during the past decade. Carditis, a dominant feature of the outbreak, was confirmed by auscultation in 53 of the patients (72 percent). An additional 14 patients were found to have mitral regurgitation by Doppler ultrasound examination, raising the total incidence of carditis to 91 percent. The children were predominantly from white (96 percent) middle-class families with above-average incomes and with ready access to medical care. There was no apparent increase in the incidence of streptococcal disease or other explanation for the marked increase in acute rheumatic fever. However, mucoid M type 18 and M type 3 group A streptococcal strains were isolated from several siblings of the patients and from schoolchildren (chosen at random) in the area. We conclude that acute rheumatic fever remains an important health problem in the United States.
Acute Disease↗
Selective radiolabeling of cell surface proteins to a high specific activity.
A procedure was developed for selective radiolabeling of membrane proteins on cells to higher specific activities than possible with available techniques. Cell surface amino groups were derivatized with 125I-(hydroxyphenyl)propionyl groups via 125I-sulfosuccinimidyl (hydroxyphenyl)propionate (125I-sulfo-SHPP). This reagent preferentially labeled membrane proteins exposed at the cell surface of erythrocytes as assessed by the degree of radiolabel incorporation into erythrocyte ghost proteins and hemoglobin. Comparison with the lactoperoxidase-[125I]iodide labeling technique revealed that 125I-sulfo-SHPP labeled cell surface proteins to a much higher specific activity and hemoglobin to a much lower specific activity. Additionally, this reagent was used for selective radiolabeling of membrane proteins on the cytoplasmic face of the plasma membrane by blocking exofacial amino groups with uniodinated sulfo-SHPP, lysing the cells, and then incubating them with 125I-sulfo-SHPP. Exclusive labeling of either side of the plasma membrane was demonstrated by the labeling of some marker proteins with well-defined spatial orientations on erythrocytes. Transmembrane proteins such as the epidermal growth factor receptor on cultured cells could also be labeled differentially from either side of the plasma membrane.
Cell Line↗
Interferon in the treatment of hairy cell leukemia.
An ongoing multicenter study has confirmed earlier observations that interferon alfa-2b (Intron A, Schering Corp., Kenilworth, NJ) is effective in the treatment of hairy cell leukemia (HCL). The 212 evaluable subjects in this trial received 2 X 10(6) U/m2 of interferon alfa-2b subcutaneously, on an outpatient basis, three times a week for 1 year. Of this group, 166 had previously undergone splenectomy, and 94 had received chemotherapy; 96% had more than 50% hairy cells in the bone marrow. Objective responses (complete, partial, and minor) were recorded in 89% of the study group and were characterized by a marked reduction in infections as well as in the need for transfusions. Complete response, defined as a normal complete blood count (CBC) and less than 5% hairy cells in the marrow, was achieved in 4% of the subjects. Partial responses (normal CBC) were attained in 74%, and minor responses (normalization of subnormal hematocrits or granulocyte or platelet counts) in 11%. Normalization of CBC usually did not occur until after 2 to 5 months of therapy. Results were comparable in splenectomized and nonsplenectomized patients. After 1 year of treatment, the responders were randomized to continue therapy for 6 months or to stop treatment and remain under observation. Preliminary findings have shown no significant differences in the incidence or time to progression of disease in the two groups, but additional data will be required before conclusions can be reached with regard to the need for maintenance therapy. Overall, the results of this study indicate that interferon alfa-2b is well tolerated, produces durable responses, and dramatically improves the quality of life in a high percentage of patients with progressive HCL.
Clinical Trials as Topic↗
Subcutaneous recombinant gamma interferon in cancer patients: toxicity, pharmacokinetics, and immunomodulatory effects.
Recombinant gamma interferon (r gamma-IFN) was administered s.c. daily to 26 patients with advanced cancer. Patients were assigned to one of six doses: 0.5, 1, 2, 4, 6, or 8 million units (MU)/m2 per d. The major toxicities were an influenza-like syndrome and fever, seen in all patients. Dose limiting toxicity occurred in 4 of 4 patients treated at 8 MU/m2. One patient with nodular poorly differentiated lymphocytic lymphoma had a mixed response, and two patients with renal cell cancer have had stabilization of disease for greater than 10 and greater than 12 months. Pharmacokinetic analysis, by radioimmunoassay, revealed mean serum r gamma-IFN concentrations up to 17 ng/ml, with maximal serum levels noted 6 to 13 h after injection. In vivo immunomodulation was assessed by natural killer (NK) cytotoxicity, monocyte activation as determined by cell surface expression of HLA-Dr, and peripheral blood mononuclear cell phenotype analysis by flow cytometry. The mean T4/T8 ratio increased from 2.1 pretreatment to 4.1 after 24 h of treatment, but returned to baseline after 7 and 28 days of treatment. Augmentation of NK function was noted after 7 days of treatment. Monocyte cell surface expression of HLA-Dr increased after 28 days of treatment at the three lowest doses. In conclusion, daily s.c. r gamma-IFN can be easily administered on an outpatient basis with minimal local skin toxicity, results in prolonged serum levels, and is associated with immunological changes of potential antitumor significance. Further study of the in vivo immunomodulatory effects induced by r gamma-IFN is indicated to help define the optimal treatment regimen.
Adult↗
The treatment of hairy cell leukemia with recombinant alfa interferon: a multi-institutional study.
This report expands on previous work with interferon alfa-2b (Intron A; Schering-Plough) in the treatment of hairy cell leukemia (HCL). Sixty-four patients were treated, with 89% achieving a response (either CR, PR or MR). The percentage of hairy cells in the peripheral blood decreased dramatically within one month and in the marrow, progressively over nine months. Normalization of blood counts was associated with a marked decline in serious infections and in the need for red cell or platelet transfusions. Preliminary results with a further group of patients confirm these satisfactory results.
Adult↗
Metal-binding protein in the Pacific oyster, Crassostrea gigas: assessment of the protein as a biochemical environmental indicator.
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Animals↗
Insulin-dependent ketosis-resistant diabetes in Ethiopia.
Anthropometric, clinical and biochemical findings were compared in 30 rural (group A), 18 urban insulin-requiring (group B) and 45 urban oral-agent-responsive (group C) newly diagnosed diabetics. Mean ages at onset were 28.3 +/- 12.0, 25.6 +/- 14.5 and 42.1 +/- 10.5 years respectively. The differences between A and C and between B and C were significant. Group A were poor and malnourished, with body mass index (BMI) 15.9 +/- 1.9 and 17.2 +/- 3.7 kg/m2 for males and females respectively, presented with a long history of classical diabetes without ketoacidosis and required insulin in modest doses. 3 of 10 cases had excess stool fat but none of 13 unselected cases had pancreatic calcification. Group C were better nourished, with BMI 22.6 +/- 2.8 and 22.4 +/- 4.5 kg/m2, and responded to oral agents. Group B, with BMI 17.2 +/- 2.6 and 18.6 +/- 3.1 kg/m2, required insulin for control but had C-peptide levels above 0.02 nmol/1 in 10 of 15 cases. Anthropometric indices for males, but not for females, were significantly lower in group A than in group B or C. There were significant differences in levels of glucose between A and B and A and C, free fatty acids between A and C and B and C, insulin between A and B and A and C and C-peptide between A and C and B and C. Of the 3 groups the rural type most closely resembled the tropical variants.
Adolescent↗