Considerations for use of platelet lysate in clinical MSC expansion

by Alexey Bersenev on November 19, 2015 · 0 comments

in cell product, mesenchymal

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Human Platelet Lysate (HPL) is widely used culture supplement for mesenchymal stromal cell (MSC) expansion. It was initially used in research labs since 1980s as growth factors-rich supplement for culture of variety of cell types. The first detailed study of HPL as alternative to Fetal Bovine Serum (FBS) for MSC culture was done by Christelle Doucet in 2005. In the last decade, HPL is increasingly used in MSC-based clinical trials.
What should be considered by developer when deciding on use of HPL? Here is my brief summary, based on recent review – Human platelet lysate: Replacing fetal bovine serum as a gold standard for human cell propagation?

In-housed made versus commercial
If you conducting small clinical study or preclinical work and located in close proximity to blood bank/ apheresis unit, you may consider to prep and test HPL in-house made HPL (auto- or allogeneic). There are many protocols you can choose from, depending on resources of your hospital.
If you’re planning to conduct large scale MSC trial with potential for commercialization, you can use allogeneic HPL products, available on the market. The industry recognizes its potential and offers variety of manufactured allogeneic HPL products on a market – Stemulate, MESEGEN, PLUS, PLTMax, Macopharma Platelet Lysate – to name a few. Are these reagents the same? No! You have to test them for your particular product and application.

Variability is HPL derivation and preparation
What is the difference between all HPL products? Well, there are a lot of differences. Let’s start from source material. The most frequent source material for HPL preparation is platelet concentrate, which could be derived from apheresis product or buffy coat or platelet-rich plasma. HPL also could be prepared from expired platelet units. Source material could be fresh, expired, irradiated and/or pathogen inactivated.
HPL preparation methods could vary significantly between manufacturers. The most frequent prep methods include: repeated freeze/thaw cycles, direct platelets activation by calcium chloride, sonification, solvent/ detergent treatment. Difference in anticoagulant as ancillary material may also have impact on final HPL product.
Yet another source of variability is pooling platelet concentrates/ units from multiple donors in manufacturing of large batches of allogeneic HPL. Manufacturers can pool as many as 50+ donors for manufacture of single HPL batch. Unlike pooled allo- HPL, autologous preparations are easier and safer. Limitations of auto- HPL preps could include comorbidities, age, restrictions for blood donation/ apheresis, absence of blood bank or specialized equipment.

Obviously, due to mentioned differences in preparation, all HPL products are different.

Safety of pooled HPL products
Since a lot of donors could be pooled for manufacturing of single batch, there is an increased risk of transmission of communicable infectious diseases. Regulatory jurisdiction, where commercial HPL are approved, plays a huge role in ensuring safety of these products. There are strict requirements for donor testing and increasing demand for pathogen inactivation in blood products. So, regulatory status of the product is very important here. You, as MSC product developer, must check and/ or request this information from manufacturers.

Culture-induced adverse effects
Cell culture-induced immunogenicity could be developed via absorption and internalization of antigens by MSC from allogeneic HPL. This possibility remains theoretical, since there are no reports on cell culture acquired immunogenicity, attributed to HPL. Impact of blood group type selection is still unclear and largely ignored by manufacturers of pooled HPL.
Yet another thing to know is genotoxicity – acquisition of chromosomal and genomic abberations in prolonged HPL-supplemented MSC culture. As of today, there are no reports on HPL-induced genomic abberations in MSC cultures.

Supply issues
According to some calculations, presented in review, supply of HPL is not as big issue as FBS:

Considering that an estimated 100 million whole blood donations are collected each year in the world [214], these figures suggest a potential for approximately 100,000e250,000 L of HPL per year from outdated platelet concentrates. Compared to the estimated volume of 500,000 L of FBS produced in 2003 [68,192], pHPL availability may be sufficient for human cell research and clinical applications.

Availability of many commercial HPL products allows to validate at least 2 suppliers for GMP manufacturing of MSC.

Lack of HPL standardization
Right now HPL production is experiencing standardization problem. Because in cell manufacturing process = product, lack of standardization may contribute to inability to transfer technology or compare results of clinical trials, performed by different groups.

However, there is a need to reach international consensus on production, quality, and safety criteria of both platelet concentrates and PRP used as raw materials and resulting HPL used as GMP- compliant supplements for ex vivo expansion of therapeutic cells for transplantation. As the use of HPL in cell therapy procedures is still emerging, it is quite conceivable that specialized human platelet derivatives will be developed in the near future both for specific therapeutic applications as for ex vivo cell expansion, thereby opening far greater potential applications than what FBS can provide.

Testing of HPL and comparison with FBS and serum-free media
The most important thing, of course, is to test HPL product for your product and application. What should be tested? Here is minimal “must” to do list:
– cell growth – proliferation/ PDL
– phenotype stability
– function check – potency (example: immunomodulatory properties)
– genotype (genomic) stability
Consider additional tests: impact of PHL on MSC differentiation, speed of senescence and potency after cryopreservation.
Ideally, you have to compare HPL product with FBS (or any other serum) and with serum-free media formulations. You better test at least 2 concentrations of HPL. There are a lot of HPL versus FBS comparison studies. For example, one of the recent studies, found lack of immunosuppressive properties MSC, cultured with HPL compare to FBS and commercial serum-free formulation.

Plelase feel free to share your experience with platelet lysate in comments! I’d especially like to hear about negative experience and HPL manufacturing hurdles and caveats.

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