A genetic strategy to control expression of human blood group antigens in red blood cells generated in vitro

Abstract

In this work focusing on the Kidd blood group system that relies on expression of hUT-B1 glycoprotein under the Jk(a) or Jk(b) antigenic configurations, we demonstrated that hematopoietic progenitors could be genetically modified to exhibit a chosen Kidd phenotype. Beyond production of atypical Kidd phenotypes, this genetic strategy could allow generation of rare blood phenotypes from hematopoietic stem cells regardless of initial donor phenotype. Potential applications for genetically modified blood include production of control samples for immunohematologic testing and for resolution of antibody detection in multiply transfused patients.

Silencing and overexpression of human blood group antigens in transfusion: Paving the way for the next steps

“…The outcome was confirmed by flow cytometry and by methods routinely used in transfusion laboratories for detection of RBCs showing Jka or Jkb antigen at their surface. This initial work suggests that there is a technological window for production of null red blood cells for use as a reference sample and that production of populations of genetically manipulated cells for transfusional purposes is a pertinent concept [31]. Another outstanding study aimed at silencing the RHAG blood group antigen was published in 2010 [32].…”

“…Genetic manipulation of CD34+ cells to obtain overexpression of the cDNA derived from SLC14A1, that codes for JK blood group determinants, prior to differentiation into RBCs in vitro heralds the use of this approach to generate new transfusion diagnostic tools [31]. In this respect, considering stem cell lines such as embryonic stem cells or induced pluripotent stem cells, as emphasized below, results coming from the literature with studies reporting overexpression of proteins in mature erythroid cells from genetically manipulated iPSC indicate that such new diagnostic tools could easily be generated using these cells as primary targets for genetic manipulation [68].…”

“…While plasmid transfection remains the most widely used technique, some studies using delivery tools derived from Adeno-Associated Virus [30], from Human Immunodeficiency Virus type-1 [7,[31][32][33], or from Murine oncoretroviruses [5,6] have been reported. Viral vector techniques are based on the use of a defective viral genome to deliver genetic material such as a RNA or DNA expression unit as a non integrated element in the target cell when using AAV derived vectors and as integrated in the genome of the target cell with retrovirus derived vectors [34][35][36].…”

“…Viral vector techniques are based on the use of a defective viral genome to deliver genetic material such as a RNA or DNA expression unit as a non integrated element in the target cell when using AAV derived vectors and as integrated in the genome of the target cell with retrovirus derived vectors [34][35][36]. A notable advantage of retroviral vectors is to facilitate genetic manipulation procedures with high gene transfer efficiency and strong transgene expression in cell lines [5][6][7] and primary erythroid cell cultures [31,32] as compared to conventional transfection techniques. Microinjection of RNA transcribed in vitro, a seldom-used technique has been described for study of the JK (a.k.a.…”

Designing peptide‐based FSL constructs to create Miltenberger kodecytes

“…Present methods capable of creating designer red blood cells with new antigenic profiles include transducing cord cells undergoing erythroid differentiation with viral vectors containing blood group alleles [1] or by attachment of antigens onto cells using KODE technology function-spacer-lipid (FSL) constructs. FSL constructs have been used in modifying embryos, spermatozoa, zebrafish, epithelial ⁄ endometrial cells, red blood cells, virions and to create quality controls systems, diagnostic panels, modify cell adhesion ⁄ interaction ⁄ separation ⁄ immobilization, and for in vitro and in vivo imaging of cells ⁄ virions [2][3][4][5][6][7][8][9][10][11].…”