Pre-clinical Trial Program of Stem Cell Preparations

In recent years, the research of stem cell-based cell therapy has developed rapidly at home and abroad.In particular, a number of clinical trials have been initiated in the fields of degenerative diseases, cirrhosis of the liver, diseases of the immune system and graft-versus-host diseases.The clinical application of stem cells is developing rapidly, and the number of clinical trials related to cell therapy is increasing exponentially.

To achieve the clinical transformation of stem cells through the IND application, sufficient preclinical studies are required to verify their safety and effectiveness. The application materials mainly include the quality control of cell products and preclinical research.

About quality control

Stem cell products In order to ensure consistent and stable quality of each generation of cells, quality control should be carried through every link of cell production process, including tissue collection, cell separation, amplification, harvesting, and product release.Each production link, need corresponding quality control method and standard.

The following lists the corresponding tests that need to be carried out at each stage to control the quality of the product. I believe that those who are familiar with the product preparation process already know the contents of these tests very well, so I won’t repeat them here.

About biosafety

What needs to be emphasized is to ensure the safety of cell products. In addition to the above quality control, biosafety is also a very important content.The Guidelines for Quality Control and Preclinical Research of Stem Cell Agents clearly stipulate that preclinical biosafety tests should include toxicity tests, abnormal immune responses, tumorigenicity, and unexpected differentiation.

Toxicity tests require the establishment of appropriate animal test models to observe various possible toxic reactions of stem cell preparations, and try to simulate the route of administration, dose and frequency of clinical applications and set reasonable groups. There is no requirement for a positive drug, and it can be set according to the product’s mechanism of action.

Abnormal immune response testing requires the detection and analysis of lymphocyte subsets and cytokines in animals to predict possible immune system abnormalities.Abnormal immune response should be evaluated in vitro and animal tests for autologous or allogeneic preparations that have been allogeneic, subcultured in vitro and specially treated. Abnormal immune response tests should not be performed for stem cells that have not been specially treated.

Oncogenicity test is to ensure that there are no factors that can immortalize cells or form tumors in the cells. It is an important risk that cell therapy products need to pay attention to. It is generally believed that tumorigenicity is related to pluripotent stem cells (such as ESC and iPSC), but corresponding tumorigenicity tests are also required before adult stem cells are used. In the past ten years of clinical trials, there has been no case of mesenchymal stem cell tumorigenicity. The requirements for the identification of tumorigenicity in the “Chinese Pharmacopoeia” are also very clear. In addition to the living cells themselves, the tumorigenicity of the cell contents (products after cell lysis) also needs to be tested.

Unexpected differentiation studies need to use appropriate cell labeling methods to study their colonization and differentiation in animals. At present, there is no research to prove that mesenchymal stem cells can differentiate in vivo.

About validity

Cellular product effectiveness is defined by the FDA as “the ability or function of a specific product to be obtained from a large amount of clinical data when tested by appropriate laboratory methods or managed in the intended manner.”The efficacy (biological potency) of a stem cell product at the preclinical stage must be considered for the appropriate indications.It is better to develop an animal model of the disease corresponding to the pathogenesis of the indications and biological indicators closely related to the therapeutic effect of MSCs, which are positively related to the therapeutic effect of the indications.

In the effectiveness research phase, the design of the experimental program needs to pay attention to two points, one is the therapeutic effect of the indication, and the other is the mechanism of action of stem cells.

The therapeutic effects of the indications are usually evaluated by the behavioral, histopathological and other testing methods of disease animal models to assess the disease process. The verification of the mechanism of action requires multiple considerations in vivo and in vitro.

Initially, in the understanding of the mechanism of stem cells, it was thought that they could migrate to damaged tissues and differentiate in vivo to replace damaged or dead cells. Unfortunately, researchers have found that stem cells can only differentiate in vitro, and differentiation in vivo has not been supported by effective evidence.

The currently recognized mechanism for stem cells to play a therapeutic role is: stem cells can migrate to the injured site and secrete cytokines, chemokines, growth factors and other substances, by recruiting tissue-specific stem cells to generate new tissues, or play an active immunomodulatory role .

The stem cell guidelines issued by the International Society for Cell Therapy (ISCT) emphasize the need to analyze the immune properties of different mesenchymal stem cell products and to test the changes in disease-related biomarkers (mRNA, surface markers, secretory proteins, etc.) of the cell products in vitro and even in vivo microenvironment.Prochymal, a bone-marrow derived mesenchymal stem cell product originally developed by Osiris, was approved in Japan for the treatment of acute graft-versus-host disease (AGVHD).One of the release elements is the determination of the level of soluble protein TNF-R1 by ELISA.

In addition to biomarkers, other characteristics of stem cells can also predict their immunosuppressive function, such as co-cultivation with PBMC (peripheral blood mononuclear cells) to detect the corresponding function of stem cell products. Donor cells are co-cultured with patient T cells, and measuring the increase in their metabolic activity can be used as a direction for mechanism research.

In addition, mitochondrial transfer is also considered to be one of the therapeutic mechanisms of stem cells. Mitochondrial transfer of mesenchymal stem cells is carried out by the release of outer vesicles or tunneled nanotubes, which has a protective effect on the resident tissue cells and regulates its inflammatory response accordingly.

If stem cell products can be successfully entered into clinical trials, in addition to ensuring the quality of the cell products, a thorough understanding of the indications and the mechanism of stem cell action should also be obtained, and a reasonable preclinical trial should be designed.Effectiveness and safety need to go hand in hand.