How to assess the probability of carcinogenesis in cell therapy

by Alexey Bersenev on July 19, 2010 · 0 comments

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this is re-blog from Stem Cell Assays

Recent editorial note by Darwin Prockop in Molecular Therapy journal highlights the problem of potential development of malignancy as a complication of cell therapy. We are coming back to this topic again, because new interesting data has come up.

As a background for this caution I’d like to remind you that in 2005 Rubio for the first time reported spontaneous human mesenchymal stem cell (MSC) transformation after standard 6-8 weeks expansion ex vivo. Since that, nearly 10 more reports were published confirming spontaneous adult stem cell transformation in long-term cell culture in mouse and human. It was the first big alarm for cell therapies based on transplantation of ex vivo expanded adult stem cells. Intriguingly, later we got a few reports showing that expanded MSC can not overcome “senescence crisis”, are stable in long-term culture and do not undergo spontaneous transformation.

So, data from different laboratories appear to be very different and conflicting. But the importance of this question is tremendous and worth more than the 100 clinical trials right now assessing the therapeutic efficacy of MSC in different conditions. This problem also highlights the weakness of assays and necessity of developing new “tumorigeneic safety criteria” for therapeutic cell products.

As a follow-up of this intriguing story, two international teams recently reported (here and here) that MSC cultures, which were previously described as spontaneously transformed, were actually cross-contaminated by malignant cells from human cancer cell lines. So, human MSC spontaneous transformation is nothing more that cell culture artifact, as noted by authors.

…we did DNA fingerprinting and/or short tandem repeat (STR) analysis comparing the normal MSC with their transformed counterparts. The analysis shows that the transformed mesenchymal stem cells (TMC) in one laboratory were cross-contaminated with human fibrosarcoma or osteosarcoma cell lines, whereas in the other laboratory cross-contamination was due to two glioma cell lines.

Now, after all these new findings, Dr. Prockop is emphasizing that we don’t have good assays to test contamination of cell product (graft) by malignant cells. More than that, we even don’t know what the probability of these events in cell culture is and how many malignant cells is enough to cause clinical disease onset after cell transplant.

Unfortunately, none of our current technologies provides a definitive test for the presence of small numbers of tumorigenic cells in the large doses required for most therapies.

The limits of our tests for tumorigenicity are severe. There are no hard data on the minimum number of tumorigenic cells necessary to produce tumors in patients, but observations with hematopoietic stem cells suggest that the number could be approximately 100 cells.

He did some calculations and came up with a number:

Therefore, a conservative estimate is that we need an assay that will guarantee that a preparation of therapeutic cells contains less than 1 tumorigenic cell per 3.5 millions.

and more on lack of assays:

Our present assays fall far short of this level of sensitivity. Classic karyotyping detects only major rearrangements of chromosomes, is subject to cultural artifacts, and samples only a small aliquot of any cell preparation. Tests for tumorigenicity in mice are meaningful only if positive, because many human tumors will not produce tumors if directly injected into immunodeficient mice.

What is the possible consequence for cell therapy? Well, we need more assays and more safety criteria now.

Therefore, the field of MSC research has rediscovered the risk of cross-contamination of cell cultures posed by malignant cells, a danger that has been known for many decades but one that still plagues the field of cancer research.

Some new assays and actions, which should eliminate these errors in future, were proposed:

First, because human errors may occur in any laboratory despite stringent working procedures, DNA fingerprinting should be compulsory for all experiments involving cell lines. Scientists should verify the cell lines in their possession and use electronic databases of authenticated DNA profiles against which they can compare their results. Second, scientific journals should require that all cell lines used in an article are verified before publication.

So, the new data showed that spontaneous transformation of expanded human MSC is more likely cell culture artifact. But it gave us a new danger – potential contamination by malignant cell lines, which should be very carefully assessed in cell product development and release.

Connotea tag: adult stem cell transformation

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also read: A delicate balance between therapeutic cell expansion and cancer

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