• Possible issues in cell preparation for cardiac cell therapy trials
    There are multiple clinical trials, assessing the efficacy of autologous bone marrow mononuclear cells in acute myocardial infarction. With a recent results of the TIME study, all 3 clinical trials (FOCUS, LateTIME and TIME), supported by US Federal Government and coordinated by Cardiovascular Cell Therapy Research Network (CCTRN), considered as failed. Everyone today is discussing the possible reasons of failures. It could be trial design, end points, statistical analysis, cell source and cell product preparation. Maybe cells simply don’t work! [...]
  • Methods and Protocols Digest – May 2013

    by Alexey Bersenev on May 22, 2013 · 0 comments

    in methods,protocols

    Methods and Protocols Digest is a collection of links to the methodological articles, freely available online. Please share your protocol with community!

    FEATURED:

    Adipose tissue-derived cells

    Mesenchymal stromal cells

    Cancer

    Neural cells

    iPS cells

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    Cells Weekly – May 19, 2013

    by Alexey Bersenev on May 19, 2013 · 0 comments

    in notes

    Cells Weekly turns one year! We started on May 20th 2012 and every week provided fresh information for you! We hope that you find this digest interesting and useful. Please leave your feedback and suggestions for the future content.

    1. Human embryonic stem cells cloned via somatic cell nuclear transfer
    It was the best paper and the most significant event of the week and, probably, of the whole year. Also it was the most highly discussed topic. Unfortunately, a lot of discussions and coverage were around “cloning hysteria“.

    @pknoepfler polled his readers on “reaction to this breakthrough” and analyzed the current human cloning ban situation in US.

    It is very much possible that premature ban of reproductive cloning will adversely affect SCNT. Therefore, under huge pressure of “cloning hysteria”, SCNT development for therapies could be halted. So, in the ethical debate about the human cloning, we always must take in account SCNT as a very promising method for generation of therapeutic cells and promote its development!

    The next big step in SCNT development is (i) reproducibility and (ii) comparison to iPS cells.

    2. Dramatic vision improvement in patient after ES cell therapy
    Advanced Cell Technology (ACT) – the only company conducting embryonic stem cell-based therapy clinical trials. This week ACT release a news about one patient with dramatic vision improvement after transplantation of their product.

    Irv Arons – independent ophthalmic industry analyst – wrote on his blog a very interesting story behind of this remarkable release and highlighted the significance:

    So, what is the significance of this development. It is significant because it shows that, for the first time, a person suffering with the dry form of AMD (90% of all those with AMD) can obtain improved vision, going from legally blind (20/400), to normal vision (20/40), good enough to obtain a driver’s license in most states. Yes, this is just one patient, and early in this clinical trial, but hope prevails.

    3. Turning point in Italian stem cell treatment scandal
    We all were watching how Italian case is unfolding. As reported by Nature News Blog, the important decision has been made this week:

    Yesterday (16 May) the Chamber’s social affairs committee unanimously passed amendments to the decree which would allow the Brescia-based Stamina Foundation, which developed the therapy, to continue administering it. However, Stamina would be required to do so within regular clinical trials, under the oversight of regulatory agencies, and using cells manufactured according to Good Manufacturing Practice (GMP). A supervisory ‘observatory’ comprising experts and patient representatives would oversee clinical trial procedures.

    Street protests continued.

    4. Poll on the ethics of payment for unproven cell therapy procedures
    I started a poll: Can patients be charged for experimental cell therapy? Please take a minute and vote. Please read description before vote!

    Create your free online surveys with SurveyMonkey , the world’s leading questionnaire tool.

    I’ll post a results later.

    5. iPS cell patents landscape in US
    iPS cell-related intellectual property is a hot topic. This week GEN posted a very interesting analysis of current iPS cell patents trends in US:

    The initial search identified approximately 230 pending and issued patents having these terms in the claims. 177 pending applications and issued patents were selected as relevant.

    6. Commercialization of cell therapies to treat stroke
    Industry analyst @JNapodano wrote a very good overview about 2 public companies – Cytomedix and Athersys, assessing their commercial products in stroke patients:

    The goal of this article is to introduce investors to the core technologies at each firm, outline each of their clinical trials currently underway in stroke, and provide a brief conclusion as to why we think cell therapy may offer hope where others have failed.

    Highly recommended!

    7. Functional hematopoietic stem cells from iPS cells
    It has been very difficult and nearly impossible to generate fully functional and serially engraftable hematopoietic stem cells (HSC) from ES and iPS cells in vitro. This week, a group of Japanese researchers has reported about derivation of engraftable HSC from iPS cell-derived teratomas:

    Here, we demonstrate a unique in vivo differentiation system yielding engraftable HSCs from mouse and human iPSCs in teratoma-bearing animals in combination with a maneuver to facilitate hematopoiesis.

    This study demonstrated the possibility of HSC differentiation from iPS cells. Because it have been done only in vivo, we still have to learn how to make it ex vivo.

    8. Returns in the Stem Cell Space – interview with Jason Kolbert
    The Life Sciences Report posted a big interview with cell therapy industry analyst Jason Kolbert. If you’re following the industry, you can find a lot of interesting things from this interview:

    A company can’t necessarily patent a cell, but the process, the method of use and, in some cases, the composition of the final product can be patented.

    When we talk about cardiovascular disease, the last thing we want to do, because the market is so large, is a highly personalized, expensive approach. Here’s where Cytori shines, because it provides basically the best of both worlds. It has an autologous process with allogeneic-like COGS.

    Clearly, the autologous companies try to maximize the allogeneic immune risks that you asserted in the previous question, and the allogeneic companies try to maximize the host comorbidity, cell-vitality argument.

    ****************
    That’s all folks. Have a fantastic week!

    bonus:

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    In the previous post we defined adipose-derived: stromal vascular fraction (SVF) versus stromal (stem) cells and highlighted difference between them. Today we will look at SVF more precisely and define its composition.

    SVF is freshly isolated heterogeneous cell fraction, which could be derived from native adipose tissue or liposuction aspirates. SVF could be derived from both – the fatty and fluid portions of liposuction aspirates after enzymatic digestion. If SVF derived from fatty portion, it’s also called PLA (processed lipoaspirates) cells.

    Basically, SVF is what remained in the pellet after removal blood and fat components. It is very crude and heterogeneous mix of multiple cell populations with different degree of maturity and function. I draw a scheme of SVF isolation and composition:

    SVF

    Based on method of adipose tissue processing, cellular composition of SVF can vary significantly. Most sources indicate that adipose-derived stromal (stem) cells represent up to 10% (2-10%) of SVF. Endothelial cells (mature and progenitors) could represents anything from 7% up to ~30% of SVF. Depending on processing, fibroblasts could represent up to 50% of SVF (Cytori data, presented at ISCT 2010). CD34+ cells are present at large number and could compose up to 63% of SVF.

    More on composition by markers:

    It has also been described that the SVF is composed of 11% CD2+ cells, 18% CD11a+ cells, 29% CD14+ cells, 49% CD31+ cells, 57% CD45+ cells, and 60% CD90+ cells (referring to ASCs and endothelial cells) [source]. Others detected a different composition of the SVF (nearly 11% CD14+ cells, ~2% CD31+ cells, ~7% CD34+, ~9% CD45+ cells, ~29% CD90+, and ~47% 146+ cells) [source].

    One more good study:

    The SVF of human adipose tissue contained: endothelial progenitors, (15.4 ± 4.8)% (mean ± standard error), pericytes (2.0 ± 1.1)%, CD146+/CD34+ transitional cells 0.5 ± 0.3, and SA-ASC (59.0 ± 10.0)% of non-hematopoietic (CD45−/CD14−/CD33−/glycophorin A−) singlet cells.

    To summarize:
    SVF composed of many mature, progenitor and stem cell types. Depending on adipose tissue processing method, the composition of SVF and relative values of each cell population can vary significantly. Adipose-derived stromal (stem) cells represent 2-10% of SVF.

    ***************
    This post is a part of series Breaking Down Fat. In this series we will talk about identification, characterization and clinical processing of potentially therapeutic cell populations from adipose tissue. We started this series in response to the growing trend of wide (mostly uncontrolled) clinical use adipose-derived cells and some controversies/ misconceptions in the field.If you would like to contribute to this series or become a sponsor, please contact us!

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    Today, a group of researchers led by Shoukhrat Mitalipov from Oregon Health & Science University published results of remarkable study – Human Embryonic Stem Cells Derived by Somatic Cell Nuclear Transfer (SCNT). As we all remember, 7 years ago, South Korean scientist Woo Suk Hwang admitted data falsification in his similar study. Few years later, after many failed attempts scientists finally make it work! This is a great news!


    So, Mitalipov’s team figured out the tricks. Now it’s time to reproduce the technique:

    The success came through minor technical tweaks. The researchers used inactivated Sendai virus (known to induce fusion of cells) to unite the egg and body cells, and an electric jolt to activate embryo development. When their first attempts produced six blastocysts but no stable cell lines, they added caffeine, which protects the egg from premature activation.

    Now we have one more way to make autologous pluripotent cells for therapies! There were some warnings in the mass media about potential human “cloning hysteria”, based on results of this study. However, Mitalipov said:

    “The embryos we produce this way did not lead to pregnancy in monkeys,” he says. “We think there is something in the manipulations to make them that make a successful pregnancy impossible.”

    Paul Knoepfler wrote on his blog:

    Overall, this paper is fascinating and a huge development, but this is a double-edged sword too.
    …the elephant in the room for this paper is the potential for future reproductive human cloning.

    I think, that the potential problem of human reproductive cloning is definitely overblown! Obviously, this is the most exciting and significant news in stem cell research of this year so far! What more interesting is a debate about medical potential of pluripotent stem cells, derived from iPS cells versus SCNT. I think, generation of therapeutic cells via SCNT is absolutely worthy to pursue! It has some advantages compare to iPS cells.

    From Nature’s piece:

    But Stojkovic, like others, awaits the results of head-to-head comparisons between iPS and SCNT cells. Some research has shown that iPS cells are not completely reprogrammed and that stem cells derived from SCNT are more like embryonic stem cells derived from in vitro fertilization. Mitalipov and Tachibana are now conducting a study to compare iPS cells and SCNT cells derived from the same donor cell. “These results,” says Daley, “will be fascinating.”

    I’ve collected some links and opinion about this great news in storify:


    Please take a poll on Knoepflelr Lab Stem Cell Blog – what’s your reaction to the human cloning news?

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    Cells Weekly – May 12, 2013

    by Alexey Bersenev May 12, 2013 notes

    Cells Weekly! is a digest of the most interesting news and events in stem cell research, cell therapy and regenerative medicine. We pick and post it every Sunday. Follow us! 1. Circulating factor able to “rejuvenate the heart” For a decade, Harvard’s investigator Amy Wagers was looking for factors circulating in the blood and able to rejuvenate aged organs and tissues. Her studies, methodologically based on conjunction of circulation young and old animals, finally brought some results. For the first [...]

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    Breaking down fat: Nomenclature of adipose-derived cells

    by Alexey Bersenev May 10, 2013 adipose

    Because of huge interest in using adipose-derived cells as “therapies”, it is important to know what is a cellular composition of this tissue. You may frequently hear a statement about fat tissue as a “richest source of stem cells” in our body. What kind of stem cells exist in adipose tissue? Can we define them? We can roughly break down fat tissue on mature adipocytes, blood and everything else. The latter frequently called “stromal vascular fraction” (SVF). SVF is enriched [...]

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    Clinical cell processing news – part 3, 2013

    by Alexey Bersenev May 8, 2013 cell product

    Clinical Cell Processing News series overviews new protocols, products and techniques for clinical-grade cell processing and manufacturing. Cell processing devices, cultureware, bioreactors, GMP-grade reagents, cell separation techniques. Follow us! 1. Validation of dry-thawing device for hematopoietic cell products (Transfusion): Progenitor cell viability and function are preserved with this dry-thawing system. The time to hematopoietic engraftment of patients after transplantation is comparable to those infused with progenitor cells thawed with the water bath technique. 2. Validation of automated system for cord [...]

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    Cells Weekly – May 5, 2013

    by Alexey Bersenev May 5, 2013 notes

    Welcome to Cells Weekly! We’re trying to navigate you in the ocean of information by picking the most interesting news and events in stem cell research, cell therapy and regenerative medicine! 1. Transplantation of tissue engineered trachea in US – Hannah Warren’s case This was a biggest mass media buzz this week. An international team, led by famous Paolo Macchiarini, has performed the first tissue engineered trachea transplant in US. You can learn more from press-conference: This case is really [...]

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    Recommendations for assessment of genetic instability of mesenchymal stromal cells

    by Alexey Bersenev May 2, 2013 mesenchymal

    We’ve written a lot about assessment of tumorigenicity risk in mesenchymal stromal cell (MSC)-based products. Recently, recommendations of the Cell Products Working Party and the Committee for Advanced Therapies (from meeting held in October 2011) have been reviewed and published online in Cytotherapy. This is very interesting and useful review, which summarizes current experience and position of regulatory agency on MSC products risk assessment. Some conclusions and recommendations from this meeting: Due to cancer cell lines contamination issues, it is [...]

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    Not Lost in Translation – Considerations for preclinical animal testing

    by Alexey Bersenev April 30, 2013 notes

    This is third post of our series “Not Lost in Translation“. Please feel free to contribute  by a post or comment! ********************** The necessity and significance of animal models in cell therapy Cell therapy is a field of medicine, which highly dependent and simply can not succeed without animal models. The necessity of animal testing is very high and only in some rare cases it could be replaced by in vitro assays. Besides proof-of-principle experiments (aimed to demonstrate that cells [...]

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