Aspects of the present disclosure relate to methods and compositions related to the preparation of immune cells, including engineered immune cells. Certain embodiments of the disclosure include compositions, cells, and methods related to engineered invariant natural killer T (iNKT) cells for off-the-shelf use for clinical therapy. The iNKT cells may be produced from hematopoietic stem progenitor cells and may be suitable for allogeneic cellular therapy because they are HLA negative. In some aspects, the cells have imaging and suicide targeting capabilities.

The present invention relates to, inter alia, an engineered cell (e.g., iPSC, IPS-derived NK, or NK cell) comprising a disrupted B2M gene and an inserted polynucleotide encoding one or more of SERPINB9, a fusion of IL15 and IL15Rα, and/or HLA-E. The engineered cell can further comprise a disrupted CIITA gene and an inserted polynucleotide encoding a CAR, wherein the CAR can be an anti-BCMA CAR or an anti-CD30 CAR. The engineered cell may further comprise a disrupted ADAM17 gene, a disrupted FAS gene, a disrupted CISH gene, and/or a disrupted REGNASE-1 gene. Methods for producing the engineered cells are also provided, and therapeutic uses of the engineered cells are also described. Guide RNA sequences targeting described target sequences are also described.

The present invention relates to a method for the production of microglia from stem cells comprising the steps of a) targeted insertion of a nucleotide sequence encoding a transcriptional regulator protein into a first genomic safe harbour site; and b) targeted insertion of the coding sequence of the transcription factor PU.1 (SEQ ID NO: 1) into a second genomic safe harbour site, wherein the gene is operably linked to an inducible promoter, which is regulated by the transcriptional regulator protein; expression of PU.1 (SEQ ID NO: 2); and culturing the stem cells received from steps a) and b) with exposure to at least one growth factor or small molecule that mimics signaling during at least one stage of embryonic development of microglia or adult microglia proliferation, differentiation or polarization. Further, the present invention relates to the microglia obtained by the methods of the present invention and various uses thereof.

The present disclosure relates to a method of culturing quiescent hematopoietic stem cells. This method involves providing a culture medium and introducing, into the culture medium, quiescent hematopoietic stem cells to culture the stem cells and maintain quiescence of the stem cells. The culture medium comprises a vacuolar-H.sup.+ adenosine triphosphate ATPase (“v-ATPase”) inhibitor. Also disclosed are methods of treating a subject for a hematological disorder, methods of culturing leukemic stem cells, and methods of enhancing the hematopoietic reconstitution ability of a population of human hematopoietic stem cells.

The present invention relates to methods and compositions for treating a hematological malignancy with an expanded hematopoietic stem cell product in combination with a chemotherapy regimen.

Aspects of the present disclosure relate to methods and compositions related to the preparation of immune cells, including engineered T cells comprising at least one exogenous γδ T cell receptor, for example one that is selected to target a specific disease or pathogen (e.g., cancer or COVID-19). The T cells may be produced from human hematopoietic stem/progenitor cells and are suitable for allogeneic cellular therapy because they do not induce graft-versus-host disease (GvHD) and resist host immune allorejection. Consequently, such cells are suitable for off-the-shelf use in clinical therapy.

The present invention relates to a method for producing induced natural killer (iNK) cells, into which a chimeric antigen receptor (CAR) gene encoding a CAR is introduced, iNK cells produced by the method, and a cell therapy composition and a pharmaceutical composition for preventing or treating cancer, comprising the iNK cells.

The method according to the present invention has the effects of producing the iNK cells, into which a CAR gene is introduced, with high efficiency through direct reprogramming from isolated cells without limiting an initial cell, and directly producing the same without a differentiation process, thereby simplifying the production process and reducing costs and time. The method according to the present invention has the effect of producing excellent NK cells having enhanced safety by directly producing NK cells from human somatic cells that are easy to obtain, without passing through induced pluripotent stem cells produced through conventional reprogramming technology. In addition, the iNK cells, into which a CAR gene is introduced, produced by the method, have an excellent cancer cell killing ability, and thus can be effectively utilized as a cell therapy composition or a pharmaceutical composition for preventing or treating cancer.

The present invention relates to, inter alia, an engineered cell (e.g., iPSC, IPS-derived NK, or NK cell) comprising a disrupted B2M gene and an inserted polynucleotide encoding one or more of SERPINB9, a fusion of IL15 and IL15Rα, and/or HLA-E. The engineered cell can further comprise a disrupted CIITA gene and an inserted polynucleotide encoding a CAR, wherein the CAR can be an anti-BCMA CAR or an anti-CD30 CAR. The engineered cell may further comprise a disrupted ADAM17 gene, a disrupted FAS gene, a disrupted CISH gene, and/or a disrupted REGNASE-1 gene. Methods for producing the engineered cells are also provided, and therapeutic uses of the engineered cells are also described. Guide RNA sequences targeting described target sequences are also described.

The present disclosure provides a method for producing red blood cells by using peripheral blood as well as the produced red blood cells.

The present invention provides a pluripotent stem cell comprising a co-expression vector in which Runx1 and Hoxa9 are of in tandem, and a T cell differentiated therefrom and application thereof. In the present invention, Pluripotent stem cells inducibly co-expressing exogenous Runx1 and Hoxa9 are successfully established by introducing an exogenous vector co-expressing Runx1 and Hoxa9 into pluripotent stem cells. The pluripotent stem cells are directionally differentiated into T-lineage progenitor cells and will be developed into T cells. The pluripotent stem cell-derived T cells obtained by the method of the present invention are not only functionally normal but also have no tumorigenic risk.