13 results about "V600E" patented technology

The invention discloses an antigen polypeptide related to BRAF gene V600E mutation and application of the antigen polypeptide. On the basis of a classical tumor neoantigen affinity prediction technology, an advanced immunogenicity prediction AI algorithm is adopted to improve the accuracy of tumor neoantigen prediction; the antigen polypeptide related to BRAF gene V600E mutation is characterized in that a replacement peptide for enhancing immunogenicity is further excavated based on a biological information screening technology of amino acid editing, the sequence of the antigen polypeptide related to BRAF gene V600E mutation is shown as any one of SEQ ID NO: 1-3, and the antigen polypeptide can remarkably activate T cells specifically aiming at BRAF mutation of a human body and enhance the killing ability of the T cells to BRAF mutation tumor cells. Therefore, the antigen polypeptide obtained by the invention has good potential of polypeptide vaccines and good clinical transformation and disease prevention prospects.

The invention provides a kit for detecting human BRAF gene V600E and TERT gene promoter mutation, and belongs to the technical field of biology. The kit comprises a primer, a probe, a 5 * PCR (Polymerase Chain Reaction)-Enhancer, a 10 * PCR Buffer, dNTP (Deoxyribonucleoside Triphosphate) Mix, MalTaq DNAPlyase, magnesium ions, DEPC (Diethylpyrocarbonate) water, a positive control and a negative control. According to the present invention, the ARMS-PCR detection technology is adopted to detect the specific gene mutation, and the target mutation site is identified by designing the specific primer and the probe, such that the detection specificity and the detection sensitivity are improved; by combining with a Taqman probe technology, the fluorescent signal change in the PCR amplification process can be monitored in real time, so that whether mutation of a BRAF V600E site, a TERT C228T site and a TERT C250T site exists or not is accurately judged; according to the method, pre-amplification of 15 cycles is adopted, then quantitative amplification of 30 cycles is adopted, and when the initial amount of the target DNA is very low, the copy number of the target DNA can be increased, so that the detection accuracy and sensitivity are further improved.

The invention provides application of an intracellular lactic acid level increasing agent in preparation of a drug for enhancing sensitivity of BRAF V600E mutant tumors to a BRAF inhibitor or in preparation of a drug for treating BRAF V600E mutant tumors resistant to the BRAF inhibitor. According to the invention, the intracellular lactic acid concentration is improved by using an intracellular lactic acid level improver, SUMOylation of BRAFV600E protein is promoted, and the BRAFV600E protein is locked in an open drug sensitive conformation; in-vivo and in-vitro experiments prove that the combination of the MCT4 inhibitor and the BRAF inhibitor (such as vimofenib) has a remarkable synergistic anti-tumor effect, drug resistance can be effectively relieved, and a new treatment strategy is provided for refractory colorectal cancer and the like.

The present application relates to a primer for quantitative detection of V600E mutation in the human BRAF gene based on digital PCR, the primer comprising the nucleotide sequences shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3. Preferably, the probe comprises a wild-type probe of the V600E gene, a mutation site of the V600E gene (c.1799_1800delinsAT), a mutation site of the V600E gene (c.1799_1800delinsAC) and / or a mutation probe of the mutation site of the V600E gene (c.1798_1799delinsAA). The present invention combines the vibration droplet generation digital PCR (dPCR) technology and, through innovative methodological optimization (artificial sequence), demonstrates significant technical advantages and clinical application value in the detection of V600E mutation in the human BRAF gene.

The application belongs to the technical field of immunochemistry, and particularly relates to an anti-BRAF-V600E recombinant rabbit monoclonal antibody and application thereof, which comprises a heavy chain variable region and a light chain variable region, the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO: 4, and the amino acid sequence of the light chain variable region is shown as SEQ ID NO: 5. The application also relates to a nucleotide sequence, a recombinant plasmid or an expression vector, a preparation method and application of the anti-BRAF-V600E recombinant rabbit monoclonal antibody in a BRAF-V600E protein detection method or device. The BRAF-V600E recombinant rabbit monoclonal antibody of the 135K5D1 clone of the application is good in specificity, strong in positive signal and the like, and is easier to score in IHC staining, and is more accurate in detection and distinction of cancer.

The invention discloses a noninvasive exosome mRNA BRAF V600E mutation Tag-PCR (Tag-Polymerase Chain Reaction) detection method, which comprises the following steps: separating an exosome sample from a body fluid; the method comprises the following steps: by taking mRNA as a template, adding an exogenous reference gene and a primer pair for detecting the exogenous reference gene, and meanwhile, carrying out a first round of PCR amplification reaction by using a Tag-ARMS primer pair for specifically enriching exosome mRNA BRAF gene V600E mutation so as to form a PCR product; a PCR product is used as a template, a system without UNG enzyme is used for amplification, a primer pair for detecting a reference gene is added, a Tag primer pair for specifically enriching and detecting V600E mutation of the human exosome mRNA BRAF gene and a probe sequence are used at the same time, and then a second round of PCR amplification reaction is carried out; and judging the mutation state of the V600E site of the BRAF gene of the mRNA of the exosome sample.

The application discloses a combination of a MEK inhibitor and a PLCbeta / PKC signal path inhibitor and application of the combination in treating BRAF(V600E) mutant tumors, and belongs to the field of biological medicines. The PLCbeta / PKC signal path inhibitor is combined with the MEK inhibitor, drug resistance can be reversed, stronger anti-BRAF(V600E) mutant tumor activity is exhibited, the growth of tumors can be more effectively inhibited, and the defects of poor treatment effect or large toxic side effects of single drug treatment are overcome. The application has a good application prospect.

The present invention relates to a set of primers for DNA amplification, which have the following sequences: 5'-CTGATAGGAAAATGAGATCTACTG-3' (SEQ ID NO: 1) 5'-GGCCAAAAATTTAATCAGTGG-3' (SEQ ID NO: 2) 5'-TCTGTAGCTAGACCAAAATCACCTATTTTCCTTTACTTACTACACCTC-3' (SEQ ID NO: 3) 5 '-AGAAATCTCGATGGAGTGGGTCCAAAATAGCCTC AATTCTTACC AT-3 ' (SEQ ID NO: 4) 5'-TCTTCATGAAGAAATATATCT-3' (SEQ ID NO: 5) 5'-CATCAGTTTGAACAGTTGT-3' (SEQ ID NO: 6). The present invention further relates to a diagnostic kit, a method for detecting a V600E mutation in the BRAF gene in a DNA sample, and to the use of said set of primers and / or of the diagnostic kit for detection of the BRAF V600E mutation in a DNA sample.

The invention belongs to the technical field of immunochemistry, and particularly relates to an anti-BRAF-V600E recombinant rabbit monoclonal antibody and application thereof.The anti-BRAF-V600E recombinant rabbit monoclonal antibody comprises a heavy chain variable region and a light chain variable region, and the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO: 4; the amino acid sequence of the light chain variable region is as shown in SEQ ID NO: 5. The invention also relates to a nucleotide sequence for coding the anti-BRAF-V600E recombinant rabbit monoclonal antibody, a recombinant plasmid or an expression vector, a preparation method of the anti-BRAF-V600E recombinant rabbit monoclonal antibody, application of the anti-BRAF-V600E recombinant rabbit monoclonal antibody in a BRAF-V600E protein detection method or device, and the like. The BRAF-V600E recombinant rabbit monoclonal antibody cloned from 135K5D1 has the characteristics of good specificity, strong positive signal and the like, so that scoring is easier in IHC staining, and cancer detection and distinguishing are more accurate.

The invention relates to a primer for quantitatively detecting V600E mutation of a human BRAF gene based on digital PCR (Polymerase Chain Reaction). The primer comprises nucleotide sequences as shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3. The invention also relates to a method for quantitatively detecting the V600E mutation of the human BRAF gene based on the digital PCR. Preferably, the probe comprises a wild type probe of the V600E gene, a mutation site (c.17991800delinsAT) of the V600E gene, a mutation site (c.17991800delinsAC) of the V600E gene, and / or a mutation probe (c.17981799delinsAA) of the mutation site of the V600E gene, and the mutation probe of the mutation site of the V600E gene can be used for detecting the mutation of the V600E gene. According to the method, a vibration microdroplet generation digital PCR (dPCR) technology is combined, and through innovative local law optimization (artificial sequence), remarkable technical advantages and clinical application value are shown in human BRAF gene V600E mutation detection.

The application discloses a primer probe composition for detecting a BRAF V600 gene mutation, a kit and application thereof. The application discloses a primer probe composition for detecting a BRAF V600 gene mutation, which comprises forward primers V600M-F and V600E-F shown in SEQ ID NO. 1 and SEQ ID NO. 2, a reverse primer V600R shown in SEQ ID NO. 3 and a detection probe V600P shown in SEQ ID NO. 4. Further disclosed are a kit containing the primer probe composition and application thereof. The primer probe composition is used for detecting a sample to be detected, has high sensitivity and specificity, can detect multiple BRAF V600 gene mutations, and can directly determine whether there is a mutation according to an amplification curve, without the need of calculating ΔCt, and is simple and direct.

This invention discloses the application of the small molecule drug PLX4032 in the preparation of a drug for the treatment of cerebral arteriovenous malformation. The small molecule drug PLX4032 of this invention can inhibit Braf... V600E Overactivation of the mutant protein Braf-MEK1 signaling pathway, thereby reducing Braf V600E The mutant protein's kinase activity, thereby preventing Braf from occurring in arteriovenous malformations of the brain. V600E The expression of mutant proteins can directly target diseased cells, resulting in good therapeutic effects.

The expression of the P-glycoprotein (P-gp) efflux transporter at the blood-brain interface impedes BBB penetrance of most small molecules. Designing efflux liabilities out of compounds can be laborious and there is no generalizable approach to transform periphery-limited agents to ones active in the CNS. A target-agnostic, prospective assessment of P-gp efflux using diverse compounds indicated a reduction in molecular size or appending a carboxylic acid that enabled evasion of P-gp efflux in cell-based experiments and in mice. Such strategy was applied to transform a periphery-limited V600EBRAF inhibitor, dabrafenib, into compounds that possess potent and selective anti-cancer activity, but now also evaded P-gp-mediated efflux. When compared to dabrafenib, the compound developed herein (everafenib) has superior BBB penetrance and superior efficacy in an intracranial mouse model of metastatic melanoma, suggesting it as a lead candidate for the treatment of melanoma metastases to the brain and gliomas with BRAF mutation.