773 results about "Circular RNA" patented technology

The invention discloses an RNA (Ribonucleic Acid) in-vitro cyclization and rolling circle translation method based on an I-type intron and an application of the RNA in-vitro cyclization and rolling circle translation method. According to the invention, a ribonucleic acid construct is obtained by splitting a specific non-complementary region of the I-type intron, and the preparation of'scarless' (traceless) circular RNA is realized through a ribozyme self-splicing principle and can be popularized to other I-type intron. By optimizing a splitting structure, high-efficiency cyclization can be realized without additionally adding GTP for catalysis under the condition of no homologous arm. Meanwhile, the original part of the ribonucleic acid construct is optimized, so that the cyclization efficiency is further improved. Particularly, a rolling circle translation (RCT) technical platform based on the CITE sequence is established, and the expression quantity of the protein or the polypeptide is greatly improved. Through verification, the cyclization and rolling circle translation method provided by the invention shows potential in vaccine and CAR treatment, and is expected to be used for clinical treatment of various diseases.

The present invention discloses a pharmaceutical composition comprising a circular RNA and a drug delivery carrier. Compared with traditional linear 1 * siRNA and annular 1 * siRNA, the number of repeated series connection of positive-sense strands is increased to include but not limited to two or more, it is accidentally found that the silence effect is remarkably enhanced, the expression level of the PCSK9 gene can be remarkably reduced, and degradation of mRNA of PCSK9 protein is mediated. The nano-particles are used for delivering oligonucleotide, so that the stability is improved, the immunogenicity is reduced, the effects of lowering cholesterol, reducing aortic plaque load and resisting atherosclerosis are improved, and the nano-particles are safe and free of obvious liver and kidney toxicity and have a wide application prospect in the aspect of preparing medicines for treating hypercholesterolemia and coronary heart disease.

Disclosed herein are compositions comprising polymeric lipid nanoparticles, and methods for preparing the same. The polymeric lipid nanoparticles can comprise RNA polynucleotides, such as circular RNA polynucleotides (circRNA), ionizable lipids (lipid) and amphiphilic polymers, thereby forming a polymeric lipid nanoparticle composition. In some embodiments, the lipid forms a complex with the circRNA (circRNA-lipid complex) that is encapsulated in the nanoparticles. In some embodiments, the circRNA-lipid complex is encapsulated in the core of the nanoparticles and the amphiphilic polymer provides a uniform shell around the circRNA-lipid complex. The present application also provides emulsions comprising the polymeric lipid nanoparticles.

Provided is a circular multimeric tandem RNA sense strand, comprising at least one sense strand sequence and at least one spacer sequence. The circular RNA is derived from an engineered parental DNA template containing all essential sequences, and sequentially comprises a first cyclization element, optionally at least one first restriction enzyme recognition sequence, at least one target sequence, optionally at least one second restriction enzyme recognition sequence, and a second cyclization element. The circular multimeric tandem RNA sense strand can bind and deliver a plurality of antisense strand RNA, increasing the binding of the sense strand and the antisense strand while utilizing the stability advantage of circular RNA.

The invention provides annular multi-tandem PCSK9 siRNA, which comprises at least one positive-sense strand sequence and at least one spacer sequence. The sense strand of the cyclic siRNA is derived from an engineered parent DNA template containing all essential sequences, comprising in the following order a first cyclization element, optionally at least one first restriction enzyme recognition sequence, at least one target sequence, optionally at least one second restriction enzyme recognition sequence, and a second cyclization element. The annular multi-tandem RNA positive-sense strand can combine and deliver a plurality of antisense strand RNAs, and the combination of the positive-sense strand and the antisense strand is increased while the stability advantage of the annular RNA is utilized.

This invention relates to recombinant nucleic acid molecule for making a circular RNA, comprising in the 5' to 3' direction: a) an internal guide sequence (IGS), b) a ribozyme, c) a first portion of an extended anticodon arm (eACA) sequence, d) gene of interest, and e) a second portion of the eACA sequence. A nucleotide in the second portion of the eACA sequence forms a wobble base pair with a nucleotide in the IGS. Recombinant nucleic acid molecules, circular RNA, and methods of producing and using circular RNA are provided.

The present disclosure relates, generally, to methods for producing, purifying, and using circular RNA from a prokaryotic system.

Provided is a pharmaceutical composition, comprising a circular RNA and a drug delivery carrier. It is surprisingly found that compared with conventional linear 1 x siRNAs and circular 1 x siRNAs, increasing the number of tandem repeats in the sense strand to two or more significantly enhances the silencing effect, significantly reduces the expression level of the PCSK9 gene, and mediates the degradation of the PCSK9 protein mRNA. The use of nano-particles for delivering oligonucleotides features improved stability, reduced immunogenicity, improved cholesterol-lowering, aortic plaque load-reducing and anti-atherosclerotic effects, good safety, no significant liver and kidney toxicity, and therefore good prospects in preparing medicaments for treating hypercholesterolemia and a coronary heart disease.

Provided is a circular concatemeric PCSK9 siRNA, comprising at least one sense strand sequence and at least one spacer sequence. The sense strand of the circular siRNA is derived from an engineered parental DNA template containing all essential sequences, which comprises, in the following order: a first cyclization element, optionally at least one first restriction enzyme recognition sequence, at least one target sequence, optionally at least one second restriction enzyme recognition sequence, and a second cyclization element. The sense strand of the circular concatemeric RNA can bind and deliver a plurality of antisense strand RNAs, enhancing the binding between the sense and antisense strands while leveraging the stability advantage of the circular RNA.

The invention relates to the technical field of biological medicines, discloses application of circPDK1 protection, and particularly relates to application of a reagent for specific detection of circPDK1 in preparation of products for detection of esophageal cancer or prognosis evaluation of esophageal cancer. According to the application of the protected circular RNA in preparation of the esophageal cancer detection product, the circular RNA is circPDK1, and the nucleotide sequence of the circular RNA is shown as SEQ ID NO: 1. Experiments prove that circPDK1 is highly expressed in esophageal cancer tissues, growth, proliferation and migration of esophageal cancer cells are promoted, occurrence of esophageal cancer is promoted, knock-down of the circular RNA can significantly inhibit growth, proliferation and migration of esophageal cancer tumors, and the average survival time of esophageal cancer patients with low circPDK1 expression is longer than that of esophageal cancer patients with high circPDK1 expression. And a new research direction is provided for esophageal cancer detection, treatment and prognosis.

According to the invention, a structural domain reaction substrate sequence consisting of Exon1 (E1), P1 and Exon2 (E2) sequences of ribozyme is mutated, and E1, P1 and E2 sequences are mutated under the condition of maintaining the structural stability, so that the Azoarcus group I intron ribozyme still has enzyme activity and can maintain the capability of forming circular RNA (Ribose Nucleic Acid). The invention discloses a flexible vector construction method for preparing circular RNA (Ribonucleic Acid) without limitation of a substrate sequence, which comprises the following steps: determining a target to-be-cyclized site sequence NNUNNNN, and segmenting the target to-be-cyclized site sequence NNUNNNN into E1: NNU and E2: NNNN; with E1 and E2 sequences as references, designing IGS sequences to respectively form complementary pairing with E1 and E2; and 5'and 3 'homologous arms, IRES, CDS and other elements are respectively added. The method provided by the invention can be used for preparing the circular RNA for any target sequence, has no residual sequence, and has relatively high cyclization efficiency. The FlexCirc cyclization system designed on the basis of Azoarcus group I intron ribozyme can form the circular RNA, the cyclization substrate sequence has the characteristic of flexible design, and the cyclization efficiency can realize a relatively high cyclization proportion.

The invention discloses a circular RNA drug sensitivity correlation identification method based on integrated multi-instance learning. The circular RNA drug sensitivity correlation identification method comprises the following steps: collecting experimental verification circular RNA and drug sensitivity correlation data; establishing a feature representation model based on a heterogeneous network and vertexes; embedding a heterogeneous graph node into the model, and extracting deep feature representation of the node; designing a meta-path instance embedding projector, and generating a plurality of meta-path instances; a circular RNA and drug sensitivity association predictor is constructed and completed; constructing an integrated heterogeneous graph network deep learning model; and outputting the meta-path instance of the circular RNA and drug pair and the attention coefficient, and carrying out interpretable analysis. According to the method, integrated learning and a deep learning model are combined, so that the reliability of the model is improved; according to the invention, interpretable analysis is carried out by utilizing the meta-path and the attention coefficient, the potential action mechanism of the circular RNA associated with the drug sensitivity can be explained, and the guidance of medical research is facilitated.

The invention relates to an intervertebral disc degenerative change analysis system based on multi-dimensional information. The system comprises a data processing module, a modifiable factor processing module, a circular RNA data processing module, a biomarker processing module, an image processing module, a treatment information processing module, a diagnostic feature anchoring module, a model generation module and a change analysis module. Anchoring is carried out on circular RNA, biomarkers, treatment information and modifiable factors through image information to form an anchoring network, and a reliable and effective basis can be provided for disease course analysis through big data and a large number of research results, so that each doctor has a targeted expectation and understanding on the disease course of a patient in the diagnosis stage; the method has the advantages that an interventional therapy scheme can be arranged in advance or the condition of a patient can be observed in real time, research results can be collected and fed back in time through big data, the information utilization rate is increased, and an information dimension integration scheme for degenerative disease recognition is created, and the accuracy of recognition results is improved.

Provided herein are circular RNA constructs comprising an IRES, and at least one expression sequence encoding binding molecule, compositions thereof, and methods of treating autoimmune disease. In particular, circular RNA comprising an IRES and a CD19 binder or a BCMA binder are provided, optionally formulated with a delivery vehicle. Precursor polynucleotides comprising an IRES, and at least one expression sequence encoding a CAR construct are also described herein.

The invention relates to the technical field of gene engineering. The invention provides a functional sequence shRNA1 (short hairpin ribonucleic acid 1) for silencing a circular RNAh-hsacirc005838, and further provides a lentiviral vector of the functional sequence shRNA1 and a construction method of the lentiviral vector. The invention also provides a lentivirus packaging titer kit containing the lentiviral vector with the functional sequence shRNA1, and also provides an application of the lentiviral vector with the functional sequence shRNA1 or the lentiviral vector with the functional sequence shRNA1 in preparation of a kit for breast cancer diagnosis. According to the invention, an interference target sequence siRNA1 is obtained by aiming at the annular target gene RNA h-hsacirc005838, then the finally obtained functional sequence shRNA1 can directly and efficiently infect cells and silence the annular RNAh-hsacirc005838 sequence, and the transfection effect is stable.

The invention relates to an application of a protein coded by circular RNA circVEGFC in an esophageal squamous cell carcinoma diagnosis product. The invention provides a protein circVEGFC-182aa translated by a circular RNA (Ribonucleic Acid) molecule circVEGFC and application of the protein circVEGFC-182aa. It is found that the expression of a circular RNA molecule circVEGFC (namely hsacirc0004496) in esophageal squamous cell carcinoma tissue is remarkably increased, the circular RNA molecule circVEGFC participates in the malignant progression of esophageal squamous cell carcinoma, the circVEGFC can encode a protein containing 182 amino acids, namely circVEGFC-182aa, the protein is subjected to endogenous expression in esophageal squamous cell carcinoma, and the invasion and migration ability of esophageal squamous cell carcinoma is promoted; the result shows that the circVEGFC plays a biological function mainly through the specific protein circVEGFC-182aa coded by the circVEGFC, so that the circVEGFC-182aa can be used as a diagnosis marker of esophageal squamous cell carcinoma to assist diagnosis.

The invention discloses a preparation method of high-purity circular RNA (Ribonucleic Acid). The preparation method comprises the following steps: constructing a cyclization skeleton sequence with self-splicing activity; the cyclization skeleton sequence comprises the following modular elements from the 5'end to the 3 'end: a T7 promoter, a 5'-terminal homologous fragment, a 3 'intron-exon fragment, a 5'-internal homologous fragment, a 5 '-terminal spacer sequence, an IRES sequence, a target fragment, a 3'-terminal spacer sequence, a 3 '-internal homologous fragment, an exon fragment-5' intron, a 3 '-terminal homologous fragment and an Xba I endonuclease site; inserting the cyclized skeleton sequence into a pUC57 vector, and carrying out plasmid amplification; an Xba I incision enzyme linearization vector is utilized to obtain an in-vitro amplification template; the method comprises the following steps: carrying out an in-vitro amplification reaction by using dsRNA-free T7RNA polymerase to obtain dsRNA-free precursor RNA; cyclizing the precursor RNA to obtain crude circular RNA; treating the crude circular RNA by utilizing poly (A) or poly (U) polymerase, and adding poly (A) or poly (U) long tails to linear RNA in the crude circular RNA; and purifying by using an affinity chromatography column to obtain the high-purity circular RNA.

The present disclosure relates to compositions, methods, processes, kits and devices for selecting, designing, preparing, manufacturing, formulating and / or using polynucleotides having internal ribosome entry site (IRES) sequences, IRES-like sequences, or combinations thereof. In particular, the present disclosure relates to compositions, methods, processes, kits and devices for selecting, designing, preparing, manufacturing, formulating and / or using cyclic polynucleotides (e.g., cyclic RNAs). The present disclosure also relates to methods of improving expression, functional stability, immunogenicity, ease of manufacture and / or half-life of therapeutic products encoded by the circular RNA.

The invention provides a non-small cell lung cancer auxiliary diagnosis composition and application and a kit thereof, and relates to the technical field of biological detection.The non-small cell lung cancer auxiliary diagnosis composition comprises a circular RNA ALG8 primer and a circular RNA CAMTA1 primer, circALG8 and circCAMTA1 can be used as markers of early non-small cell lung cancer liquid biopsy, and the non-small cell lung cancer auxiliary diagnosis composition can be applied to diagnosis of non-small cell lung cancer. According to the present invention, with the application of the circALG8 primer and the circCAMTA1 primer, the plasma circALG8 and circCAMTA1 combined diagnosis model can be successfully constructed, the NSCLC patient and the healthy control can be effectively distinguished, the area under the curve is up to 0.8987 when the circALG8 primer and the circCAMTA1 primer are combined to identify the efficiency of the early NSCLC, the identification result has extremely high accuracy, the diagnosis accuracy is significantly improved, the sensitivity reaches 91.78%, and the specificity reaches 75.53%.

The present disclosure relates, generally, to compositions and methods for producing, purifying, and using circular RNA.

The invention relates to a method for engineering transformation of cells by using scar-free annular mRNA and application of the method. Specifically, the present inventors make circular mRNA for in vitro and / or in vivo protein production based on a cyclization technique of a secondary structure of circular RNA. The annular mRNA does not contain any extra sequence except the target sequence, and is an accurate and efficient mRNA cyclization strategy, and the generated scar-free annular mRNA has the advantages of good stability, long half-life period, high protein expression total amount and the like, can be used for CAR and / or TCR related cell therapy, is especially suitable for in-vivo CAR generation cell therapy, and has a good application prospect. The compound has wide application in the field of clinical treatment of tumors, autoimmune diseases, infectious diseases and the like.

The invention discloses a construction method, a prediction system and a prediction method of a circular RNA (Ribonucleic Acid) and disease association prediction model based on a shared unit, and association prediction is realized through the constructed circular RNA and disease association prediction model. Comprising the following steps: constructing a circular RNA meta-path network, a circular RNA similarity network, a disease meta-path network and a disease similarity network based on a known associated data set, and respectively carrying out feature extraction on the networks to obtain circular RNA similarity features, circular RNA meta-path features, disease similarity features and disease meta-path features; the similarity features and the meta-path features are simultaneously input to a sharing unit; similarity features output by the sharing unit are input into a multi-layer perceptron to carry out circular RNA and disease association prediction and update model parameters. A sharing unit is designed for the model, and the prediction performance of the model is improved by constructing a similarity network and a meta-path network of circular RNA and diseases and capturing potential cross-view information in a multi-view feature fusion process.

The application discloses a kind of based on shared unit's circular RNA and the construction method of disease association prediction model, prediction system and prediction method, and the association prediction is realized by the circular RNA and the disease association prediction model constructed, comprising: based on known association dataset constructs circular RNA meta-path network, circular RNA similarity network, disease meta-path network and disease similarity network, the feature extraction of the aforementioned network respectively obtains circular RNA similarity feature, circular RNA meta-path feature, disease similarity feature and disease meta-path feature;Similarity feature and meta-path feature are simultaneously input into shared unit;The similarity feature output by shared unit is input into multilayer perception machine to carry out the association prediction of circular RNA and disease and update model parameter.Share unit is designed for model, and the similarity network and meta-path network of circular RNA and disease are constructed, and potential cross-view information is captured in the process of multi-view feature fusion, so as to improve the prediction performance of model.

The invention discloses an application of circular RNA (Ribonucleic Acid) hsacirc0008581 as a biomarker for auxiliary diagnosis of esophageal cancer and / or prognosis prediction of esophageal cancer patients. The circular RNA hsacirc0008581 is highly expressed in esophageal cancer tumor tissues, the highly expressed hsacirc0008581 can significantly promote the increase of malignant phenotypes such as invasion, migration, metastasis and the like of esophageal cancer cells, and meanwhile, the high expression level of the hsacirc0008581 is significantly related to the shorter total lifetime of esophageal cancer patients, that is, the hsacirc0008581 can be used as an independent risk prognosis prediction factor of the esophageal cancer patients. Therefore, the hsacirc0008581 can be used as a novel biomarker for effectively assisting in diagnosing the esophageal cancer and / or predicting the metastasis risk and / or the total lifetime of the esophageal cancer patient and a novel target for treating the esophageal cancer.

The invention provides a novel IRES sequence suitable for circular RNA and a variant and an untranslated region sequence of the IRES sequence. The invention provides a circular RNA drug containing the IRES sequence, the untranslated region sequence and the protein coding sequence. The circular RNA drug platform has efficient protein translation ability (especially in immune cells), simplified sequence length and compatibility to various protein sequences, and the drug based on the circular RNA platform shows a good treatment effect in an animal tumor model.

The invention provides a nano preparation for promoting tissue repair and application thereof, the nano preparation comprises circular RNA for coding growth factors, circular RNA for coding cytokines and a nano carrier, and the preparation method comprises the following steps: preparation of a lipid carrier material, preparation of a solution of the circular RNA for coding the growth factors and the cytokines, and microfluidic mixing. Belongs to the technical field of biological medicine. The nano preparation prepared by the invention can simultaneously and accurately express and release growth factors and cell factors in cells, plays a role for a long time, regulates an immune environment, inhibits inflammation and promotes wound tissue repair, has the characteristics of no toxicity, high safety, convenience in use, simplicity in preparation and excellent effect, and has clinical application value.

The present application provides a circular RNA (circRNA) that encodes an antigen polypeptide of an SARS-CoV-2 variant. In some embodiments, the present application provides a circRNA vaccine against an SARS-CoV-2 variant, such as a Delta or Aumike variant. In some embodiments, a circRNA vaccine comprises a circRNA comprising a nucleic acid sequence encoding an antigenic polypeptide comprising a spike (S) protein of the SARS-CoV-2 variant, or a fragment thereof. Also provided are methods of treating or preventing SARS-CoV-2 infection using the circRNA disclosed herein, or a composition thereof.

Provided are methods of preparing a circular RNA, comprising providing a template DNA, wherein the template DNA comprises a sequence encoding a precursor RNA, in a reaction solution to allow synthesis of precursor RNA by in vitro transcription of the template DNA and allowing the precursor RNA to self-splice, thereby producing a circular RNA, wherein the in vitro transcription of the template DNA and the self-splicing (i.e., circularization) of the precursor RNA are carried out in the same reaction solution under the same reaction conditions (e.g., the same reaction temperature). The method can be carried out in a single reaction vessel and does not require a step of purifying the precursor RNA before allowing the precursor RNA to self-splice.

Provided is a circular RNA encoding CARs and the use thereof to create immune cells that target specific diseases, e. g., lymphoma, multiple myeloma and leukemia and auntoimmune diseases, such as systemic lupus erythematousus, lupus nephritis and myasthenia gravis.

The invention discloses application of annular RNACirc3426 as a diagnostic marker and a therapeutic target of colorectal cancer. Experiments show that the Circ3426 is highly expressed in colorectal cancer cells and patient serum, and prove that the Circ3426 has good stability, and an ROC curve shows that the Circ3426 has good capacity of diagnosing colorectal cancer. Meanwhile, it is found that the molecule can promote migration and invasion ability of colorectal cancer cells, nude mouse subcutaneous tumor formation experiments prove that knock-down Circ3426 inhibits growth of tumors, and it is indicated that the molecule can serve as a treatment target of colorectal cancer. The invention provides a new diagnostic marker and a treatment target for colorectal cancer, and provides a new thought and direction for CRC diagnosis and treatment.