Efficient Bone Marrow Transduction by Gene Transfer with Allogeneic Umbilical Cord Blood Serum and Plasma: An Implication for Clinical Trials

Abstract

Low in vivo transduction efficiency and safety concerns have been hurdles for effective hematopoietic stem cell (HSC) gene therapy. Here, we investigate whether the safety and efficiency of retroviral gene transfer into HSCs can be improved by using human allogeneic umbilical cord blood (UCB)-derived supplements instead of fetal bovine serum (FBS). When CD34(+) cells were cultured ex vivo in UCB-derived serum (CBS) or plasma (CBP), comparable or higher maintenance of HSCs was observed than in FBS and serum-free substitution medium (SFM) as assessed by the frequency of positive engraftment and the level of engraftment in NOD/SCID mice after transplantation of cultured cells. CBS and CBP also exhibited higher level stabilization of retroviral particles than SFM during in vitro culture of retrovirus pseudotyped with gibbon ape leukemia virus or vesicular stomatitis virus glycoprotein. Retroviral gene transfer into CD34(+) cells performed with CBS or CBP resulted in increased gene transfer into CD34(+) cells and increased transduction of reconstituted bone marrow cells compared to transfers with SFM or FBS. The increased transduction of bone marrow cells was associated with a larger number of transduced progenitors in the recipient mice. Significant oligoclonality in the transduced progenitors, as determined by ligation-mediated polymerase chain reaction, suggested efficient retroviral targeting of multiple HSCs in the CBS- or CBP-supplemented media. Combined, our results show that allogeneic UCB-derived serum or plasma is a safe and easily accessible serum supplement that can support efficient retroviral gene transfer into HSCs for the clinical-grade manipulation of HSCs.

An Effective and Safe Supplement for Stem Cells Expansion Ex Vivo: Cord Blood Serum

“…4A). These results revealed that cord blood serum contained abundant growth factors to maintain the proliferation potential of hPDMSCs over passages and to obtain large amounts of hPDMSCs ex vivo within a short time as others reported (15,25,36).…”

“…So it is necessary to find a replacement for FBS in stem cell expansion throughout long-term culturing ex vivo. These include autologous or allogeneic human serum, human plasma, CBS, and human platelet derivatives (7,15,17,20,22,24,25,27,28,33,36). Of note, CBS is safe serum medium that carries no risk of xenogeneic antigen and has lower immunogenicity.…”

“…Of note, CBS is safe serum medium that carries no risk of xenogeneic antigen and has lower immunogenicity. In addition, more distinct stimulating factors in CBS can contribute to the unique cellular characteristics of human MSCs; it is suggested that CBS may similarly provide a unique microenvironment for ex vivo expansion of MSCs (15,25,36).…”

Undifferentiated hematopoietic cells are characterized by a genome-wide undermethylation dip around the transcription start site and a hierarchical epigenetic plasticity

“…22 Human hematopoietic cells were similarly cultured ex vivo in serum-free substitution media containing 20% BIT as described. 25 Each culture was supplemented with growth factor cocktails containing 100 ng/mL human Flt3 ligand (R&D Systems), 100 ng/mL human stem cell factor (R&D Systems), and 20 ng/mL human interleukin-3, granulocyte colony-stimulating factor, and human interleukin-6 (R&D Systems).…”

Plasma‐derived from human umbilical cord blood restores ovarian function and improves serum reproductive hormones levels in mice with premature ovarian insufficiency (POI) through cytokines and growth factors

“…It is clear from previous research that CBP was shown to be richer for cytokines and growth factors compared to ABP and PRP (Ehrhart et al, 2018; Rhéaume et al, 2022). This approach has yielded notable successes in the treatment of neurodegenerative diseases, experimental autoimmune encephalomyelitis and hematopoietic stem cell gene therapy (Ehrhart et al, 2018; Kim et al, 2020; Moon et al, 2008). Despite these advancements, investigations into the impact of CBP derived from hUCB on POI remain notably scarce.…”