RBC manufacture without high EPO costs

The need for a reliable supply of O-negative (universal donor) red blood cells has driven ex vivo RBC manufacture closer to reality in recent years. However, the necessity for erythropoietin (EPO) in the cell culture to promote lineage maturation has remained a significant cost burden. New work from Stanford and UCSF aims to overcome this.

The research utilises protein and genome engineering to create a synthetic EPO receptor (SynEPOR) on HPSCs that is induced by a small molecule rather than by EPO. This is achieved with equivalent performance to EPO but with a 500-fold reagent cost reduction.

In flow cytometric analysis, to determine the differentiation towards fully matured RBCs, DRAQ5™ was employed to show extent of enucleation. Far-red fluorescing DRAQ5™ has been confidently employed in this way for two decades (Fraser, 2005).

The cell-engineering concept described in this work should be transferable to other clinically desirable cells and cell behaviours, to similarly put phenotypic differentiation under the control of small molecule inductions (or suppressions) to achieve a desired terminal cell functionality.

References:

Shah, A.P., Majeti K.R., et al. "Engineering synthetic signaling receptors to enable erythropoietin-free erythropoiesis." Nature Communications 16.1 (2025): 1140.

Fraser, S.T., Isern J., and Baron, M.H. "Characterization of Circulating Primitive Erythroid Cells during Embryogenesis Using a Human ε-Globin-GFP Transgenic Mouse Model." Blood 106.11 (2005): 3611.

Exciting New Macrophage Cell Therapy Option

New research published in Nature Communications offers an exciting new cell therapy option using allogeneic macrophage drug conjugates (MDC) to better attack solid tumors, in an effort to overcome some of the limitations experienced by other cell-based therapies.  

The work was led by scientists at Cellis AG, a European biotechnology company with focus in cell-based oncology therapy based in Zurich, Warsaw University of Life Sciences and collaborators in Berlin and Edinburgh.

The methodology relies upon a novel mechanism for the preferential in vitro uptake of human heavy chain ferritin (HFt) by human monocyte-derived macrophages (hMDM). This novel clathrin-mediated endocytosis mechanism, in addition to the well-understood transferrin receptor 1 (TfR1/CD71), predominantly utilised a Class A Scavenger Receptor - MSR-1. Both receptors are highly expressed on macrophages. The resulting efficient loading of macrophages was shown not to be cytotoxic to the cells.

Further, central to the therapeutic strategy was the ability to pre-load HFt with small molecule drugs at useful drug/protein ratios. The resulting HFt-drug conjugates were found to be a highly stable resource to load hMDM.

hMDM were found to be the most suitable cell type, importantly of allogeneic origin and which were highly amenable to cryopreservation.

The resulting allogeneic macrophage drug conjugates (MDC) were then tested for their ability to target cancer cells and to determine the mechanism.  MDCs targeted tumor cells and cell killing relied upon Hft-drug conjugate transfer from MDC to tumour cells by direct cell-cell contact by a process termed TRAIN (TRAnsfer of Iron proteiN). A wide range of solid tumour cell lines were tested, with low side-effects, highly targeted to the tumour cells and demonstrating good levels of apoptosis. In a variety of in vivo models, xenograft tumours shrank, cell proliferation was reduced and animals maintained weight.

MDCs were also combined with stand-alone therapeutic agents e.g. checkpoint inhibitors, to significant effect.

DRAQ7™ was used in a variety of assays to determine both unwanted and desired cytotoxicities. DRAQ7™ has ideal properties for development of cell therapies, in cross-platform cell-based assays - time-lapse cell-cell killing assays, product quality assurance analysis and more. It is a chemically-defined reagent manufactured following the ISO 9001:2015 quality system.

Reference:
Taciak B, Bialasek M, Kubiak M, et al. Harnessing macrophage-drug conjugates for allogeneic cell-based therapy of solid tumors via the TRAIN mechanism. Nature Communications ( 2025) 16:1327. https://doi.org/10.1038/s41467-025-56637-9

DRAQ5 for FISH counterstaining intracellular pathogens

In this new work to determine the role of purines in innate immunity a group led by scientists at UCSD used C elegans orthologs of enzymes human purine salvage pathway. 

In one aspect of this work they also treated HUVECs with concentrations of deoxyadenosine with and without anti-IFN antibody and then infected the cells with the intracellular parasite E. intestinalis.

DRAQ5 was used as counterstain for FISH on the HUVEC cells. The analysis contained E. intenstinalis 16S rRNA probe tagged with Quasar 570 chromophore (DsRed channel), Calcofluor White to detect bacterial cell wall (DAPI channel)

DRAQ5 to label nuclei (Cy5 channel).  The data were then used to quantify any significant impact on the infection load in the cells according to the dose of deoxyadenosine, as a measure of enhanced innate immunity.

Due to Calcofluor White's emission being in the DAPI channel DRAQ5 was chosen as DNA counterstain for these FISH experiments, showing its ideal utility for experiments to study intracellular parasitic infections since there is also great value in combining the FISH labeling with Calcafluor White's ability to label the prokaryotic cell wall. 

Reference:
Wernet, N.D., Tecle, E., Sarmiento, M.B., Kuo, C.-J., Chhan, C.B., Baick,
I., Batachari, L.E., Franklin, L., Herneisen, A., Bhabha, G., Ekiert, D.C., Hanna-Rose, W., Troemel, E.R.
Adenosine deaminase and deoxyadenosine regulate intracellular immune response in C. elegans.
ISCIENCE (2025), DOI: https://doi.org/10.1016/j.isci.2025.111950.