A DNA plasmid useful for diagnostic and therapeutic gene therapy is disclosed. Improvements to gene therapy methods known in the art are provided to ensure cancer-targeting, high efficacy, and long durability of expression. The DNA plasmid is combined with compositions of polymeric nanoparticles for non-viral gene therapy to treat cancer, including hepatocellular carcinoma and prostate cancer.

The presently disclosed subject matter provides compositions, methods, and kits for transfecting adipose-derived mesenchymal stem cells (AMSCs) in freshly extracted adipose tissue using nanoparticles comprising biodegradable polymers self-assembled with nucleic acid molecules. The presently disclosed subject matter also provides methods for treating a neurological disease in a patient in need thereof, the method comprising administering the AMSCs transfected with the nucleic acid molecules to the patient, wherein the nucleic acid molecules encode one or more bioactive molecules functional in the treatment of a neurological disease, particularly wherein the neurological disease is a brain tumor.

Provided herein are methods and compositions for a suicide gene approach comprising an expression vector comprising a cell cycle-dependent promoter driving the expression of a suicide gene. Also provided herein are methods to render proliferative cells sensitive to a prodrug after transplantation but avoids expression of the suicide gene in post-mitotic cells, such as neurons.

Modified macrophage immune cells are provided for treatment of cancer and other diseases. In particular said macrophages express chimeric antigen receptors (CAR). The single chain variable fragment (scFv) may be directed against thymidine kinase 1 (TK1) or hypoxanthine guanine phosphoribosyltransferase (HPRT). The signaling domain may be derived from a Toll-like receptor (TLR).

Provided is a method for knocking in a gene of interest to a cell. The genome of the cell contains a negative selectable marker, e.g., a thymidine kinase gene flanked by a pair of recombinase recognition sites (RRS), e.g., attP. The method involves introducing into the cell a targeting construct that contains a gene of interest flanked by a second pair of RRS, e.g., attB. The targeting construct also contains in the vector backbone a negative selectable marker, e.g., thymidine kinase gene. When a recombinase recognizing the RRS is expressed, the recombination events between the two pairs of RRS result in the site-specific integration of the gene of interest in the genome of the cell. Upon selection based on the negative selectable marker, the parental cells, cells with undesired integration, e.g., random integration, or the integration of the vector backbone are removed.

The disclosure is directed to a gene transfer vector which comprises (i) all or part of a viral genome and (ii) a suicide gene flanked by unique cloning sequences. The disclosure also is directed to a system for producing an adenoviral vector comprising a destination vector comprising (i) all or part of an adenoviral genome and (ii) a suicide gene flanked by unique cloning sequences; a transgene flanked by unique cloning sequences; and (c) reagents for Gibson DNA Assembly (GDA). A method of producing an adenoviral vector using the aforementioned system also is provided.

A method is disclosed for decreasing or retarding an increase in the size of a localized or metastatic tumor by using a combination of an immune stimulating cytotoxic gene therapy and immune-checkpoint modulating agent, in conjunction with other therapies, including radiation therapy, chemotherapy, surgery, and immunotherapies.

The present invention provides a genetically modified NK cell line prepared by transducing host NK cell line with a gene construction encoding a cancer antigen-specific chimeric antigen receptor (CAR) comprising a FLT3-specific monoclonal antibody or a functional fragment thereof, a transmembrane domain, and a CD3ζ domain of a T-cell receptor for more efficient immunotherapy of acute myeloid leukemia, and use thereof, for more efficient immunotherapy of acute myeloid leukemia.

A method for predicting patient survival comprises determining a level of STK1 (serum thymidine kinase 1) material in a body sample from a patient diagnosed with prostate cancer using an antibody or a fragment thereof specifically binding to a serum form of human TK1. The method also comprises predicting survival of the patient based on the determined level of STK1 material in the body sample.

The presently disclosed subject matter provides compositions, methods, and kits for transfecting adipose-derived mesenchymal stem cells (AMSCs) in freshly extracted adipose tissue using nanoparticles comprising biodegradable polymers self-assembled with nucleic acid molecules. The presently disclosed subject matter also provides methods for treating a neurological disease in a patient in need thereof, the method comprising administering the AMSCs transfected with the nucleic acid molecules to the patient, wherein the nucleic acid molecules encode one or more bioactive molecules functional in the treatment of a neurological disease, particularly wherein the neurological disease is a brain tumor.