Forcing Sugars on the Surfaces of Cord Blood Cell Increases Their Engraftment

When a child or adult needs new bone marrow, a bone marrow transplant from a donor is usually the only way to save their life. Without properly functioning bone marrow, the patient’s blood cells will die off, and there will be too few red blood cells to ferry oxygen to tissues or white blood cells to fight off infections.

An alternative to bone marrow from a bone marrow donor if umbilical cord blood. Umbilical cord blood does not require the rigorous tissue matching that bone marrow requires because the blood making stem cells from cord blood are immature and not as likely to cause tissue rejection reactions.. However, umbilical cord blood cells suffer from two drawbacks: low numbers of stem cells in cord blood and poor engraftment efficiencies.

Fortunately, some progress has been made at expanding blood-making stem cells from umbilical cord blood, and it is likely that such technologies might be ready for common use in the future. As to the poor engraftment efficiencies, a new paper in the journal Blood from the laboratory of Elizabeth J. Shpall at the University of Texas MD Anderson Cancer Center, in Houston, Texas reports a new way to increase cord blood stem cells engraftment efficiencies.

As previously discusses, delayed engraftment is one of the major limitations of cord blood transplantation (CBT). Delayed engraftment seems to be due to the diminished ability of the cord blood stem cells to home to the bone marrow. How are cells channeled to the bone marrow? A protein receptor called P- and E-selectins is expressed on the surfaces of bone marrow blood vessels. Cells that can bind these selectin receptors will pass from the circulation to the bone marrow. Thus binding selectin receptors is kind of like having the “password” for the bone marrow.

What does it take to bind the selectin proteins? Selectins bind to specific sugars that have been attached to proteins. These sugars are called “fucose” sugars. As it turns out, cord blood stem cells do not express robust levels of these fucosylated proteins. Could increasing the levels of fucosylated proteins on the surfaces of cord blood stem cells increase their engraftment? Shpall and her colleagues tested this hypothesis in patients with blood-based cancers.

Patients with blood cancers had their cancer-producing bone marrow stem cells destroyed with drugs and radiation. Then these same patients had their bone marrows refurbished with two units of umbilical cord blood. However, these cells in these cord blood units were treated with the enzyme fucosyltransferase-VI and guanosine diphosphate fucose for 30 minutes before transplantation. This treatment should have increased the content of fucosylated proteins on the surfaces of cells in the hope of enhancing their interaction with Selectin receptors on the surfaces of bone marrow capillaries.

The results of 22 patients enrolled in the trial were then compared with those for 31 historical controls who had undergone double unmanipulated CBT. There was a clear decrease in the length of time it took for cells to engraft into the bone marrow.  For example, the median time to neutrophil (a type of white blood cell) engraftment was 17 days (range 12-34) compared to 26 days (range, 11-48) for controls (P=0.0023). Platelet (a cell used in blood clotting) engraftment was also improved: median 35 days (range, 18-100) compared to 45 days (range, 27-120) for controls (P=0.0520).  These are significant differences.

These findings support show that treating cord blood cells with a rather inexpensive cocktail of enzymes for a short period of time before transplantation is a clinically feasible means to improve engraftment efficiency of CBT.  This is a small study.  Therefore, these data, though very hopeful, must be confirmed with larger studies.

Turning Stem Cells into Drug Factories

Wouldn’t it be nice to have cells that express the right molecules at the right place and the right time to augment or even initiate healing?

Researchers at the Brigham and Women’s Hospital and Harvard Stem Cell Institute have inserted modified messenger RNA to induce mesenchymal stem cells to produce adhesive proteins  (PSGL-1)and secrete interleukin-10, a molecule that suppresses inflammation. When injected into the bloodstream of mice, these modified stem cells home to the right location, stick to that site, and secrete interleukin-10 (IL-10) to suppress inflammation.

Improving MSC therapeutic potential viamRNA transfection with homing ligands and immunomodulatory factors. Illustration of (A) mRNA-engineered MSCs that express a combination of homing ligands (PSGL-1 and SLeX) and an immunomodulatory factor (IL-10), and (B) targeting mRNA-engineered MSCs to site of inflammation.
Improving MSC therapeutic potential viamRNA transfection with homing ligands and immunomodulatory factors. Illustration of (A) mRNA-engineered MSCs that express a combination of homing ligands (PSGL-1 and SLeX) and an immunomodulatory factor (IL-10), and (B) targeting mRNA-engineered MSCs to site of inflammation.

Jeffrey Karp, Harvard Stem Cell Institute principal faculty member and leader of this research, said this about this work: “If you think of a cell as a drug factory, what we’re doing is targeting cell-based, drug factories to damaged tissues, where the cells can produce drugs at high enough levels to have a therapeutic effect.”

Karp’s paper reports a proof-of-principle study has piqued the interest of several biotechnology companies, since it has the potential to target biological drug to disease sites. While ranked as the top sellers in the drug industry, biological drugs are still challenging to use. Karp’s approach might improve the clinical applications of biological drugs and improve the somewhat mixed results of clinical trials with mesenchymal stem cells.

Mesenchymal stem cells (MSCs) have emerged as one of the favorite sources for stem cell therapies. The attractiveness of MSCs largely lies with their availability, since they are found in bone marrow, fat, liver, muscle, and many other places. Secondly, MSCs can be grown in culture for a limited period of time without a great deal of difficulty. Third, MSCs tend to be ignored by the immune system when injected. For these reasons, MSCs have been used in many clinical trials, and they appear to be quite safe to use.

To genetically modify MSCs, Karp and his co-workers made chemically modified messenger RNAs (mRNAs) whose bases differed slightly from natural mRNAs. These chemical modifications did not affect the recognition of the mRNA by the protein synthesis machinery of the MSCs, but did affect the recognition of these mRNAs by those enzymes that degrade mRNAs. Therefore, these synthetic mRNAs are very long-lived and the transfected cells end up making the proteins encoded by these mRNAs for a very long time. RNA transfection does not modify the genome of the host cells, and this makes it a very safe procedure, since the engineered cells will express the desired protein for some time, but not indefinitely.

The mRNAs introduced into the cultured MSCs included mRNAs that encode the IL-10 protein, which is cytokine that suppresses inflammation, the PSGL-1 protein, a cell-surface protein that sticks to the P-and E-selectin receptors, and the Fut7 gene product.  FUT7 encodes an enzyme called fucosyltransferase 7, which adds a sugar called “fucose” to the PSGL-1 protein and without this sugar, PSGL-1 cannot bind to the selectins.  Selectins are stored by cells and during inflammation, they are sent to the cell surface where they can bind cells and keep them there to mediate inflammation.  By expressing PSGL-1 in the MSCs, Karp and his group hoped to that the engineered MSCs would bind to the surfaces of blood vessels and not be washed out.


Oren Levy, lead author of this paper, said, “This opens the door to thinking of messenger RNA transfection of cell populations as next generation therapeutics in the clinic, as they get around some of the delivery challenges that have been encountered with biological agents.”

A problem that constantly troubles clinical trials that use MSCs is that they are rapidly cleared from the bloodstream within a few hours or days after they are introduced. The Harvard team showed that rapid targeting of MSCs to inflamed tissue produced a therapeutic effect despite rapid clearance of the MSCs.

Karp and his colleagues would like to extend the lifespan of these cells in the bloodstream and they are presently experimenting with new synthetic mRNAs that encode pro-survival factors.

“We’ve interested to explore the platform nature of this approach and see what potential limitations it may have or how far we can actually push it. Potentially we can simultaneously deliver proteins that have synergistic therapeutic impacts,” said Weian Zhao, another author of this paper.