71 results about "Dihydrofolic acid" patented technology

The invention discloses a Chinese hamster ovary (CHO) genetic engineering cell line for performing high-level secretory expression on AcAPc2 and relates to a CHO cell line for performing high-level secretory expression on the AcAPc2. The invention aims to solve the problems that the expression level of an exogenous gene product expressed by using CHO is low at present and the industrialized production cannot be realized. The CHO genetic engineering cell line for performing high-level secretory expression on the AcAPc2 is obtained by the following step of: screening dihydrofolate reductase-deficient CHO cells which are taken as host cells, and amplifying to obtain the cell line for performing stable and high-level expression on the AcAPc2. The CHO cell line can perform high-level expression on the AcAPc2; the expression level can be up to 10mg / L.72h; and the CHO cell line can be industrially produced. The cell line is used for resisting blood coagulation and treating tumor, septicaemiaand the like.

Protein Fragment Complementation Assays (PCA) are done in plant material using enzyme fragment constructs, for example, dihydrofolate reductase (DHFR) fragment constructs. Plant material is transformed with at least two different constructs that form different products capable of interacting to reconstitute enzymatic activity in the plant material. Detection of the activity can be done using a substrate for the enzyme in the culture medium which, when reacted with the enzyme, is converted to a detectable product. One embodiment uses a substrate that can be enzymatically converted to a detectable fluorescent product. An inducer such as rapamycin or salicylic acid can be added to the culture medium to increase the level of detectable product.

Compounds of formula (I) or (II) are dihydrofolate reductase inhibitors, useful for the treatment of, for example, cell proliferative diseases:wherein A and D are independently —CHR7— or —NR7—; E and G are independently ═CR7— or ═N—; each R6 independently represents hydrogen or —OR7; R7 is hydrogen or C1-C6 alkyl; R1 is a carboxylic acid group (—COOH), or an ester group which is hydrolysable by one or more intracellular carboxylesterase enzymes to a carboxylic acid group; R2 is the side chain of a natural or non-natural alpha amino acid which does not contain a carboxyl, or carboxyl ester group; Y is a bond, —C(═O)—, —S(═O)2—, —C(═O)NR3—, —C(═S)—NR3, —C(═NH)NR3 or —S(═O)2NR3— wherein R3 is hydrogen or optionally substituted C1-C6 alkyl; L1 is a divalent radical of formula -(Alk1)m(Q)n(Alk2)p- wherein m, n and p are independently 0 or 1, and Q, Alk1 and Alk2 are as defined in the claims; X1 represents a bond; —C(═O); or —S(═O)2—; —NR4C(═O)—, —C(═O)NR4—, —NR4C(═O)NR5—, —NR4S(═O)2—, or —S(═O)2NR4— wherein R4 and R5 are independently hydrogen or optionally substituted C1-C6 alkyl; and z is 0 or 1.

A Chinese hamster ovary genetic engineering cell line capable of efficiently expressing Ancylostoma caninumanticoagulant peptide 5 (AcAP5) in secretion mode relates to a Chinese hamster ovary (CHO) cell line capable of efficiently expressing the AcAP5 in secretion mode. The cell line resolves the problem that at present, expression amount is low by using CHO to express exogenous gene products, and requirements of industrial production cannot be met. The Chinese hamster ovary genetic engineering cell line for the efficient secretion expression AcAP5 utilizes dihydrofolic acid reductase defect type Chinese hamster ovary cells as host cells, and the cell line capable of stably and efficiently expressing AcAP5 is obtained after the host cells are screened and amplified. The CHO cell line can efficiently express the AcAP5, can be produced in industrialization mode and is used for resisting blood coagulation and thrombus. Furthermore, expression amount can reach to 12 mg / L / 72h.

The present disclosure provides compounds of Formula I:or a pharmaceutically acceptable salt thereof, wherein R5, R6 and Z are as described herein. The disclosure also provides pharmaceutical compositions thereof; and methods for inhibiting DHFR activity; and methods for treating cell proliferative diseases, autoimmune disease, inflammatory disease or bacterial, fungal or parasitic infection by administering a compound of Formula I.

A process for the large-scale chemoenzymatic production of (6S)-5-methyl-5,6,7,8-tetrahydrofolic acid, also known as (6S)-5-methylTHFA, the process comprising the steps of: (1) reducing folic acid (FA) so as to yield dihydrofolic acid (DHFA); (2) stereoselectively reducing DHFA with dihydrofolate reductase (DHFR) in the presence of NADP / NADPH, glucose and GluDH so as to yield (6S)-THFA; (3) converting (6S)-THFA to (6S)-5-methylTHFA; and (4) isolating (6S)-5-methylTHFA.

The invention discloses derivatives and salts of damino dihydrotriazine, and a preparation method, a composition and application thereof. According to the invention, the preparation method of the damino dihydrotriazine derivative and the damino dihydrotriazine salt can be realized by adopting a method I or a method II, wherein the method I includes the step of obtaining a general formula I compound prepared through the reaction between a general formula IV compound and a general formula V compound, while the method II includes the step of mixing a general formula VIII compound with a general formula II compound under an acidic condition, and obtaining the compound shown in the general formula I through a cyclization reaction of the mixture. The invention also provides application of derivatives and salts of the damino dihydrotriazine in preparation of human dihydrofolate reductase inhibitors, preventing and curing drugs for tumor or bacterial infection diseases. The invention further provides a drug composition, which comprises an effective amount of the derivatives and / or salts of the damino dihydrotriazine, as well as pharmaceutically acceptable carriers. According to the invention, spiro heterocyclic ring derivatives of the damino dihydrotriazine have an excellent inhibitory activity on human dihydrofolate reductase, tumor cells and bacteria.

A novel screening assay for identifying therapeutic agents and their cellular targets is described. The assay is useful in developing new antibacterial, antifungal, antiparasitic and anti cancer therapeutics. New inhibitors of dihydrofolate reductase (DHFR) have been identified and their cellular target confirmed using the assay of the present invention. Methods of treating diseases that benefit from an inhibition of DHFR are also described.

The invention discloses a recombinant CHO cell strain capable of stably expressing GPC3 and an application thereof. The preservation number of the CHO cell strain is CGMCC No.18881, the GPC3 is encoded by a cDNA sequence shown in the formula of SEQ ID No. 1, an antigen produced by the recombinant CHO cell strain is used for preparing a GPC3 monoclonal antibody, and the recombinant CHO cell strainis used for preparing a GPC3 antibody. The folate reductase defect type CHO cells are lack of endogenous dihydrofolate reductase, when an expression vector containing a dihydrofolate reductase gene and a target gene is co-transfected, the dihydrofolate reductase gene and the target gene are generally integrated at the same site of a host chromosome, and then in the presence of a competitive inhibitor-methotrexate of dihydrofolate reductase, the increasing of the copy number of the dhfr gene is driven to achieve the purpose of increasing the expression quantity of the target gene, so that a stable cell line for stably and efficiently expressing GPC3 is constructed, and the technical blank is filled.

The compositions and methods described herein disclose the design, synthesis and testing of compounds that act as inhibitors of DHFR. The basic scaffold of these inhibitors includes a 2,4-diaminopyrimidine ring with a propargyl linker to another substituted aryl, bicyclo or heteroaryl ring. These DHFR inhibitors are potent and selective for many different pathogenic organisms, including the DHFR enzyme from bacteria such as Bacillus anthracis and methicillin-resistant Staphylococcus aureus, fungi such as Candida glabrata, Candida albicans and Cryptococcus neoformans and protozoa such as Cryptosporidium hominis and Toxoplasma gondii. These compounds and other similar compounds are also potent against the mammalian enzyme and may be useful as anti-cancer therapeutics.

The invention aims at providing an application of dihydrofolate synthetase in detection of sulphonamide residue and a rapid sulphonamide detection method constructed by utilizing the dihydrofolate synthetase. The dihydrofolate synthetase capable of detecting the sulphonamide residue is an amino acid sequence shown in a sequence SEQ ID NO.1. Compared with the traditional detection method, the dihydrofolate synthetase has the advantages that firstly the dihydrofolate synthetase is wide in source, protein fragments just containing a dihydrofolate synthetase core sequence can be applied, and the dihydrofolate synthetase can be rapidly and stably obtained by adopting a genetic engineering method; and secondly, a rapid detection method for all the sulphonamides containing a p-aminobenzene sulphonamide parent core structure can be constructed by utilizing the new application of the dihydrofolate synthetase.

Novel medicinal compositions containing anti-Fas antibody which are useful as preventives or remedies for autoimmune diseases or rheumatoid arthritis. More particularly, medicinal compositions containing as the active ingredients antihuman Fas antibody having an activity of inducing apoptosis and a compound having an antagonism to folic acid or an effect of inhibiting dihydrofolate reductase. By using these medicinal compositions, the content of anti-Fas antibody can be reduced in preventives or remedies for autoimmune diseases or rheumatoid arthritis containing anti-Fas antibody. Thus, the possibility of the appearance of the tolerance in the patient's body due to the expression of an antibody against the anti-Fas antibody, etc., which makes it possible to provide excellent preventives or remedies usable in long-term administration.

The invention discloses a method for preparing a receptor-ligand binding sulfonamide antibiotic protein. The method comprises the following steps: firstly preparing a prokaryotic expression vector, and then isolating and purifying Escherichia coli strains to obtain dihydrofolate synthase; with a genome of Escherichia coli as a template, taking the obtained dihydrofolate synthetase gene fragment and cloning into the prokaryotic expression vector to obtain a cloning vector, and extracting recombinant plasmids from the cloning vector and the prokaryotic expression vector; culturing the recombinant plasmids and Escherichia coli in a culture medium to obtain a positive clone strain, and then continuously culturing the positive clone strain in the culture medium and centrifuging to obtain a thallus; concentrating the thallus and polyethylene glycol octylphenyl ether, and centrifuging to obtain a protein; and finally purifying the protein. According to the method for preparing the receptor-ligand binding sulfonamide antibiotic protein provided by the invention, a rapid detection product produced under the action of high binding force has sensitivity higher than that of the same product produced by an antibody binding principle.

The invention relates to inhibitors of dihydrofolate reductase and pharmaceutical preparations thereof. The invention further relates to methods of treatment of parasitic infections, such as T. gondii, T. cruzi, P. falciparum, T. brucei, or L. major infections, using the novel inhibitors of the invention.

The invention relates to a reagent (kit) for measuring formaldehyde by using an enzyme-multiplied method, an enzyme colorimetric method and an enzyme coupling method as well as a method for measuring the concentration of the formaldehyde, and composition and ingredients of the reagent, belonging to the technical field of food / environmental test. The reagent (kit) comprises the main ingredients of buffer solution, coenzyme, adenosine triphosphoric acid, dihydrofolic acid, glyceric aldehyde-3-phosphoric acid, formaldehyde dehydrogenase, formic acid-dihydrofolic acid ligase, glyceric aldehyde-3-glycerol phosphate dehydrogenase and stabilizing agent. The concentration of the formaldehyde is measured by mixing a sample and the reagent according to a certain volume ratio to carry out a series of enzymatic reaction, placing the reactant under an ultraviolet / visible light analyzer and detecting the ascending degree of absorbance at a dominant wavelength of 340nm.