Provided is a bacteria for targeting tumors and treating cancer, a drug delivery composition, and methods of using same for treating cancer.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

The present disclosure relates to T cells capable of co-expressing T cell receptors (“TCR”) together with CD8 polypeptides and the use thereof in adoptive cellular therapy. The present disclosure further provides for modified CD8 sequences, vectors, and associated methods thereof.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

Provided is a nucleic acid system introduced into a bacterial strain to generate a genetically engineered bacterial strain that grows in solid tumors but does not grow in non-tumor tissues, the nucleic acid system comprising: a first DNA fragment that encodes a toxin gene that expresses a toxin that kills the genetically engineered bacterial strain; a second DNA fragment that encodes an antidote gene that expresses an antidote that negates the toxin; a first promotor that controls transcription of the antidote gene, such that glucose represses the transcription of the antidote gene; and a first constitutive promoter that causes constitutive expression of the toxin gene; wherein the second DNA fragment is transcribed in the solid tumors but not transcribed in the non-tumor tissues.

Provided is a nucleic acid system introduced into a bacterial strain to generate a genetically engineered bacterial strain that grows in solid tumors but does not grow in non-tumor tissues, the nucleic acid system comprising: a first DNA fragment that encodes a toxin gene that expresses a toxin that kills the genetically engineered bacterial strain; a second DNA fragment that encodes an antidote gene that expresses an antidote that negates the toxin; a first promotor that controls transcription of the antidote gene, such that glucose represses the transcription of the antidote gene; and a first constitutive promoter that causes constitutive expression of the toxin gene; wherein the second DNA fragment is transcribed in the solid tumors but not transcribed in the non-tumor tissues.

The present invention provides a circular DNA vector, which may be used to introduce specific a mutation in a target region of a host cell. The present invention further provides methods using the circular DNA vector for generating engineered host cells. The circular DNA vector and methods are useful for studying gene functions and generating cells producing recombinant gene products.