Methods and compositions are provided for editing the genome of a cell without the use of an exogenously supplied nuclease. Aspects of the methods include contacting a cell with a targeting vector comprising nucleic acid sequence to be integrated into the target locus, where the cell is not also contacted with a nuclease. In addition, reagents, devices and kits thereof that find use in practicing the subject methods are provided.

The present invention provides a genetically-modified T cell comprising in its genome a modified human T cell receptor alpha gene. The modified T cell receptor alpha gene comprises an exogenous sequence of interest inserted into an intron within the T cell receptor alpha gene that is positioned 5′ upstream of TRAC exon 1. The exogenous sequence of interest can comprise an exogenous splice acceptor site and/or a poly A signal, which disrupts expression of the T cell receptor alpha subunit. The sequence of interest can also include a coding sequence for a polypeptide, such as a chimeric antigen receptor. Additionally, the endogenous splice donor site and the endogenous splice acceptor site flanking the intron are unmodified and/or remain functional. The invention further provides compositions and methods for producing the genetically-modified cell, and populations of the cell, and methods for the treatment of a disease, such as cancer, using such cells.

Compositions and methods are provided for integrating one or more genes of interest into cellular DNA without substantially disrupting the expression of the gene at the locus of integration, i.e., the target locus. These compositions and methods are useful in any in vitro or in vivo application in which it is desirable to express a gene of interest in the same spatially and temporally restricted pattern as that of a gene at a target locus while maintaining the expression of the gene at the target locus, for example, to treat disease, in the production of genetically modified organisms in agriculture, in the large scale production of proteins by cells for therapeutic, diagnostic, or research purposes, in the induction of PS cells for therapeutic, diagnostic, or research purposes, in biological research, etc. Reagents, devices and kits thereof that find use in practicing the subject methods are also provided.

The invention relates to constructs, vectors, relative host cells and pharmaceutical compositions which allow an effective gene therapy, in particular of genes larger than 5 Kb by using an improved hybrid dual recombinant AAV vector system.

The present invention provides assays for profiling two or more polypeptides in live cells. In some embodiments, the invention provides multicistronic reporter vectors, acceptor cells for receiving multicistronic reporter vectors, and multireporter cells. Methods of making multicistronic reporter vectors, acceptor cells for receiving multicistronic reporter vectors, and multireporter cells are provided. Libraries and kits comprising multicistronic reporter vectors, acceptor cells for receiving multicistronic reporter vectors, and multireporter cells are provided. Methods of profiling/assaying the multireporter cells and multireporter cell libraries are provided.

The invention provides compositions and methods for reprogramming minimal volumes of mononuclear cells. In particular aspects, the invention provides methods and compositions for reprogramming minimal volumes of umbilical cord blood obtained from cord blood segments from cryopreserved cord blood segments.

The present invention relates to a non-viral minicircle vector expressing a SOX gene, a stem cell into which the vector is introduced, a pharmaceutical composition for preventing or treating a cartilage disease, including the stem cell, and a method for constructing the vector. The transformation of mesenchymal stem cells with MC/SOX-Trio or MC/SOX-Duo, which is a non-viral minicircle vector according to the present invention, can completely exclude the necessity of expensive growth factors that have been indispensably used in inducing the differentiation of mesenchymal stem cells into chondrocytes. Accordingly, the mesenchymal stem cells transformed therewith, when implanted in vivo, can differentiate into chondrocytes by themselves, and thus have an advantage capable of simplifying the existing complicated steps of culturing cells to induce differentiation and then transplanting the cells. Further, unlike existing vector systems in which antibiotic-resistant genes and other bacteria-derived exogenous genes are simultaneously transferred to cells even after transformation, the vector of the present invention minimizes transfer of unnecessary genes into target cells by allowing two or three SOX genes necessary only for differentiation into chondrocytes to be regulated under one promoter, and thus can be utilized as a non-viral vector system in the most advantageous form for use in clinical application of stem cell-gene therapeutic agents.

The present disclosure provides nucleic acid constructs for the treatment of cancer, comprising a cancer-specific promoter and one or more therapeutic genes.

The present invention relates to the field of genetic engineering, in particular, to a transposon-based transfection kit suitable for transfection of primary cells, such as T cells, comprising mRNA encoding a transposase, or reagents for generating mRNA encoding said transposase, as well as minicircle DNA comprising the transposon. The invention also relates to a nucleic acid, preferably, a DNA minicircle, comprising a transposon, wherein the transposon encodes a protein and at least one miRNA, wherein the sequences encoding the miRNA are located in an intron and expression of the protein and the miRNA is regulated by the same promoter. The invention also provides a population of cells obtainable with the method of the invention. Methods of transfection are also provided, as well as medical use, e.g. in immunotherapy, in particular, in adoptive T cell therapy or T cell receptor (TCR) or chimeric antigen receptor (CAR) gene therapy.

Provided herein are methods and compositions for treating cancer in an individual comprising administering to the individual an effective amount of at least one CD 122/CD 132 agonist, at least one immune checkpoint inhibitor and a viral composition comprising one or more viruses engineered to overexpress a tumor suppressor gene and/or an adenoviral death protein. Also provided herein are methods and compositions for treating cancer in an individual comprising administering to the individual an effective amount of at least one oncolytic viral composition and at least one CD122/CD132 agonist and at least one immune checkpoint inhibitor. Also provided herein are methods of enhancing anti-tumor efficacy by administering the agents described above in combination with other cancer therapies. In highly aggressive forms of cancer, known to be generally resistant to immune therapies, these treatments unexpectedly resulted in complete tumor remissions and curative outcomes.