The preparation of a semisynthetic tritiated insulin with a specific radioactivity of up to 20 Curies per millimole.
Ontology highlight
ABSTRACT:
SUBMITTER: Halban PA
PROVIDER: S-EPMC1172350 | biostudies-other | 1975 Nov
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Semisynthetic [3H]insulin was stored under various conditions for up to 180 days and the stability of the insulin under these conditions was assessed. A sample that had been stored for 180 days was repurified and shown to be degraded at the same rate as native insulin by perfused mouse livers, even at low physiological concentrations. After perfusion, intact insulin could be separated from degradation products, and the radioactivity associated with the insulin fraction could be used to determine the percentage degradation. The initial rate of degradation of insulin was a linear function of concentration over the range 360pM-1.9nM.
| S-EPMC1164249 | biostudies-other
Project description:We have prepared and characterized semisynthetic [GlyA1-3H]insulin. The preparation was carried out at specific radioactivities ranging from 1Ci/mmol to 44Ci/mmol. The largest quantity prepared in any one synthesis was 3.5 mCi. Chemical degradation showed that the label was in its expected position in the molecule. The semisynthetic product behaved authentically on reversed-phase h.p.l.c. and radioimmunoassay. It gave the expected profiles of biological activity as regards depression of blood sugar concentration in rats and stimulation of conversion of glucose into lipid in isolated rat fat-cells. We discuss some applications for which this tracer would be particularly suited. An expanded version of this paper, containing full experimental details of the semisynthesis and characterization of [GlyA1-3H]insulin, has been deposited as Supplementary Publication SUP 50129 (30 pages) at the British Library (Lending Division), Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1985) 225, 5.
| S-EPMC1152758 | biostudies-other
Project description:1. Conditions favouring the condensation of o-phenylenediamine with alpha-oxo acids to form substituted quinoxalines have been investigated. The best yields (85-100%) are obtained by incubation of the reagents in 2m-hydrochloric acid. Substitution of acetic acid for hydrochloric acid, or of halogen-substituted 1,2-diaminobenzenes for phenylenediamine, increases the initial rate of reaction, but not the yield of quinoxalines. 2. The quinoxaline derivatives are insoluble in water and can be readily purified. They are very suitable for determination of the specific radioactivities of radioactively labelled alpha-oxo acids because they are colourless, soluble in dioxan-based scintillators, and their strong absorption of u.v. light facilitates their chemical determination. As little as 1 nmol can be accurately determined by means of the blue fluorescence produced by absorption of light at 340-360nm. 3. Optimum conditions for preparation, purification and separation by paper and thin-layer chromatography are described.
| S-EPMC1179581 | biostudies-other
Project description:Lignans, neolignans, norlignans and norneolignans constitute a large class of phenolic natural compounds. 9-Norlignans, here defined to contain a β⁻β' bond between the two phenylpropanoid units and to lack carbon number 9 from the parent lignan structure, are the most rarely occurring compounds within this class of natural compounds. We present here an overview of the structure, occurrence and biological activity of thirty-five 9-norlignans reported in the literature to date. In addition, we report the semisynthetic preparation of sixteen 9-norlignans using the natural lignan hydroxymatairesinol obtained from spruce knots, as starting material. 9-Norlignans are shown to exist in different species and to have various biological activities, and they may therefore serve as lead compounds for example for the development of anticancer agents. Hydroxymatairesinol is shown to be a readily available starting material for the preparation of norlignans of the imperanene, vitrofolal and noralashinol family.
| S-EPMC6358742 | biostudies-literature
Project description:BackgroundThe employment of well characterized test samples prepared from authenticated, high quality medicinal plant materials is key to reproducible herbal research. The present study aims to demonstrate a quality assurance program covering the acquisition, botanical validation, chemical standardization and good manufacturing practices (GMP) production of IBS-20, a 20-herb Chinese herbal formula under study as a potential agent for the treatment of irritable bowel syndrome.MethodsPurity and contaminant tests for the presence of toxic metals, pesticide residues, mycotoxins and microorganisms were performed. Qualitative chemical fingerprint analysis and quantitation of marker compounds of the herbs, as well as that of the IBS-20 formula was carried out with high-performance liquid chromatography (HPLC). Extraction and manufacture of the 20-herb formula were carried out under GMP. Chemical standardization was performed with liquid chromatography-mass spectrometry (LC-MS) analysis. Stability of the formula was monitored with HPLC in real time.ResultsQuality component herbs, purchased from a GMP supplier were botanically and chemically authenticated and quantitative HPLC profiles (fingerprints) of each component herb and of the composite formula were established. An aqueous extract of the mixture of the 20 herbs was prepared and formulated into IBS-20, which was chemically standardized by LC-MS, with 20 chemical compounds serving as reference markers. The stability of the formula was monitored and shown to be stable at room temperature.ConclusionA quality assurance program has been developed for the preparation of a standardized 20-herb formulation for use in the clinical studies for the treatment of irritable bowel syndrome (IBS). The procedures developed in the present study will serve as a protocol for other poly-herbal Chinese medicine studies.
| S-EPMC2845568 | biostudies-literature
Project description:First-generation cysteine-based site-specific antibody-drug conjugates (ADCs) are limited to one drug per cysteine. However, certain applications require a high drug to antibody ratio (DAR), such as when low-potency payloads are used. Higher drug load can be achieved using classical cysteine conjugation methods, but these result in heterogeneity, suboptimal efficacy and pharmacokinetics. Here, we describe the design, synthesis and validation of heterobifunctional linkers that can be used for the preparation of ADCs with a DAR of two, three and four in a site-specific manner per single cysteine conjugation site, resulting in site-specific ADCs with a DAR of four, six and eight. The designed linkers carry a sulfhydryl-specific iodoacetyl reactive group, and multiple cyclic diene moieties which can efficiently react with maleimide-carrying payloads through the Diels-Alder reaction. As a proof of concept, we synthesized site-specific DAR four, six and eight ADCs carrying tubulysin (AZ13601508) using engineered antibodies with a cysteine inserted after position 239 in the antibody CH2 domain. We evaluated and compared the in vitro cytotoxicity of ADCs obtained via the site-specific platform described herein, with ADCs prepared using classical cysteine conjugation. Our data validated a novel cysteine-based conjugation platform for the preparation of site-specific ADCs with high drug load for therapeutic applications.
| S-EPMC7555909 | biostudies-literature
Project description:The B-cell receptor (BCR) is a key player of the adaptive immune system. It is a unique part of immunoglobulin (Ig) molecules expressed on the surface of B cells. In case of many B-cell lymphomas, the tumor cells express a tumor-specific and functionally active BCR, also known as idiotype. Utilizing the idiotype as target for lymphoma therapy has emerged to be demanding since the idiotype differs from patient to patient. Previous studies have shown that shark-derived antibody domains (vNARs) isolated from a semi-synthetic CDR3-randomized library allow for the rapid generation of anti-idiotype binders. In this study, we evaluated the potential of generating patient-specific binders against the idiotype of lymphomas. To this end, the BCRs of three different lymphoma cell lines SUP-B8, Daudi, and IM-9 were identified, the variable domains were reformatted and the resulting monoclonal antibodies produced. The SUP-B8 BCR served as antigen in fluorescence-activated cell sorting (FACS)-based screening of the yeast-displayed vNAR libraries which resulted after three rounds of screening in the enrichment of antigen-binding vNARs. Five vNARs were expressed as Fc fusion proteins and consequently analyzed for their binding to soluble antigen using biolayer interferometry (BLI) revealing binding constants in the lower single-digit nanomolar range. These variants showed specific binding to the parental SUP-B8 cell line confirming a similar folding of the recombinantly expressed proteins compared with the native cell surface-presented BCR. First initial experiments to utilize the generated vNAR-Fc variants for BCR-clustering to induce apoptosis or ADCC/ADCP did not result in a significant decrease of cell viability. Here, we report an alternative approach for a personalized B-cell lymphoma therapy based on the construction of vNAR-Fc antibody-drug conjugates to enable specific killing of malignant B cells, which may widen the therapeutic window for B-cell lymphoma therapy.
| S-EPMC7726437 | biostudies-literature
Project description:BackgroundType II diabetes (T2D) is preceded by prolonged insulin resistance and relative insulin deficiency incompletely captured by glucose metabolism parameters, high-density lipoprotein (HDL) cholesterol and triglycerides.ObjectiveWhether lipoprotein insulin resistance (LPIR) score, a metabolomic marker, is associated with incident diabetes and improves risk reclassification over traditional markers on extended follow-up.MethodsAmong 25,925 nondiabetic women aged 45 years or older, LPIR was measured by nuclear magnetic resonance spectroscopy as a weighted score of very low density lipoprotein, low-density lipoprotein, and HDL particle sizes, and their subsets concentrations. We run adjusted cox regression models for LPIR with incident T2D (20.4 years median follow-up).ResultsAdjusting for demographics, body mass index, life style factors, blood pressure, and T2D family history, the LPIR hazard ratio for T2D (hazard ratio [HR] per standard deviation, 95% confidence interval) was 1.95 (1.85, 2.06). Further adjusting for HbA1c, C-reactive protein, triglycerides, HDL and low-density lipoprotein cholesterol, LPIR HR was attenuated to 1.41 (1.31, 1.53) and had the strongest association with T2D after HbA1C in mutually adjusted models. The association persisted even in those with optimal clinical profiles, adjusted HR per standard deviation 1.91 (1.17, 3.13). In participants deemed at intermediate T2D risk by the Framingham Offspring T2D score, LPIR led to a net reclassification of 0.145 (0.117, 0.175).ConclusionIn middle-aged or older healthy women followed prospectively for over 20 years, LPIR was robustly associated with incident T2D, including among those with an optimal clinical metabolic profile. LPIR improved T2D risk classification and may guide early and targeted prevention strategies.
| S-EPMC5644504 | biostudies-literature
Project description:The insulin receptor (INSR) binds insulin to promote body growth and maintain normal blood glucose levels. While it is known that steroid hormones such as estrogen and 20-hydroxyecdysone counteract insulin function, the molecular mechanisms responsible for this attenuation remain unclear. In the present study, using the agricultural pest lepidopteran Helicoverpa armigera as a model, we proposed that the steroid hormone 20-hydroxyecdysone (20E) induces dephosphorylation of INSR to counteract insulin function. We observed high expression and phosphorylation of INSR during larval feeding stages that decreased during metamorphosis. Insulin upregulated INSR expression and phosphorylation, whereas 20E repressed INSR expression and induced INSR dephosphorylation in vivo. Protein tyrosine phosphatase 1B (PTP1B, encoded by Ptpn1) dephosphorylated INSR in vivo. PTEN (phosphatase and tensin homolog deleted on chromosome 10) was critical for 20E-induced INSR dephosphorylation by maintaining the transcription factor Forkhead box O (FoxO) in the nucleus, where FoxO promoted Ptpn1 expression and repressed Insr expression. Knockdown of Ptpn1 using RNA interference maintained INSR phosphorylation, increased 20E production, and accelerated pupation. RNA interference of Insr in larvae repressed larval growth, decreased 20E production, delayed pupation, and accumulated hemolymph glucose levels. Taken together, these results suggest that a high 20E titer counteracts the insulin pathway by dephosphorylating INSR to stop larval growth and accumulate glucose in the hemolymph.
| S-EPMC7949120 | biostudies-literature
Project description:The process of neurodegeneration in Parkinson's Disease is intimately associated with the aggregation of the protein ?-synuclein into toxic oligomers and fibrils. Interestingly, many of these protein aggregates are found to be post-translationally modified by ubiquitin at several different lysine residues. However, the inability to generate homogeneously ubiquitin modified ?-synuclein at each site has prevented the understanding of the specific biochemical consequences. We have used protein semisynthesis to generate nine site-specifically ubiquitin modified ?-synuclein derivatives and have demonstrated that different ubiquitination sites have differential effects on ?-synuclein aggregation.
| S-EPMC3315850 | biostudies-literature