How does HSCT work?

I have been asking myself about the non-myeablative HSCT and wondering how does it work?

It is often a combination of cyclophophamide and anti-thymocyte globulin.

Cyclophosphamide is a drug that can kill anything that divides and so it will hit activated T cells but it is also pretty good at depleting B cells, because many of them have a high natural turnover. This causes the CD34+ stem cells to mobilize from the bone marrow into the blood, where they are harvested. They then get anti-thymocyte globulin. I thought that this would therefore target T cells, so to squeeze the memory B cell idea into the mix the cyclophosphamide would have to kill memory B cells.

Indeed in a paper the reduction was by about 70% CD19 B cell compared to about 25% for T cells. Switched memory B cells depleted by about 40% and unswitched by about 60%. Memory CD4 cells were down by about 20% and CD8 memory were not affected.

Anti-thymocyte globulin (ATG) is an infusion of horse or rabbit-derived antibodies against human T cells, which is used in the prevention and treatment of acute rejection in organ transplantation and therapy of aplastic anemia. I saw Dr Burt from Chicago and he said to me the anti-thymocyte globulin depletes B cells. This came as a bit of a shock but thinking about it, it makes perfect sense as T and B cells share loads of molecules that can be targeted by the antibody.

To get HSCT to optimally work we need it to clear out the immune system from the brain as it is clear it does not always do this.

Roll P, Muhammad K, Stuhler G, Grigoleit U, Einsele H, Tony HP. Effect of ATG-F on B-cell reconstitution after hematopoietic stem cell transplantation. Eur J Haematol. 2015 Dec;95(6):514-23.

Mondria T1, Lamers CHte Boekhorst PAGratama JWHintzen RQ Bone-marrow transplantation fails to halt intrathecal lymphocyte activation in multiple sclerosis.J Neurol Neurosurg Psychiatry. 2008 Sep;79(9):1013-5. doi: 10.1136/jnnp.2007.133520. Epub 2008 Jan 25.

.BACKGROUND:Given the presumed key role for autoreactive lymphocytes in multiple sclerosis (MS), treatment strategies have been developed to ablate lymphocyte activity. Intrathecal lymphocyte activation can be measured by CSF-soluble(s)CD27.

OBJECTIVE:To determine the effect of maximum whole-body immune ablation on two different markers that detect lymphocyte activation in CSF-oligoclonal IgG bands and levels of CSF-sCD27.
DESIGN, SETTING AND PATIENTS: The study quantified sCD27 levels and assessed the presence of oligoclonal IgG bands in CSF samples of secondary progressive patients with MS treated by autologous bone-marrow transplantation. In eight individuals, CSF was taken before and 6-9 months after conditioning. CSF-sCD27 levels were compared with other MS and non-inflammatory neurological disease controls. Regarding the effect of stem-cell transplantation on CSF oligoclonal bands, the study analysed pooled data of this and four other international studies on stem-cell transplantation in MS.
RESULTS:CSF-sCD27 was significantly lower after the extremely immunoablative protocol. However, levels remained elevated compared with non-inflammatory controls and stayed within the range observed in other MS controls. The joint analysis of CSF oligoclonal bands demonstrated persistence of this immune abnormality in 88% of the reported cases (n = 34).
CONCLUSIONS:The persistence of CSF lymphocyte activation markers sCD27 and intrathecal oligoclonal IgG bands after maximum immunoablative treatment indicates that complete eradication of activated lymphocytes from the CNS has not been established. This is paralleled by disease progression observed in several studies on the effect of stem-cell transplantation in MS

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  • Muraro et al in Laurence Turka's lab published in Journal of Clinical Investigation 2018 looked at sequencing of the TCR beta chain from T cells isolated pre and post HSCT from blood and found that CD4s had a tendency to expand from new clonaltypes, while the CD8s were expanded more from pre-existing memory.

    In a supplemental figure, they also discuss the presence of HLA-A2 and HLA-B8 EBV reactive CD8 clones as being present (looks like reactive against EBNA3 and BMLF; latent and lytic at least).

    The CD8 therapy in clinical trials by Atara is directed against expanding lytic specific CD8s with the suggestion that an overabundance of latent specific CD8s causes too much EBV reinfection of B cells to occur and they end up exhausted, with runaway amplification of the B cell memory pool.

    Thinking of an integrated B and T cell response- if you suggest that the memory B cell pool does not require CD4 help to stimulate damage/relapse, then is it the resetting of the CD8 lineage towards more lytic antigens (which they don't specifically say whether that has happened here) which again is better at controlling the EBV memory population post HSCT? Or could it also be that you are eliminating the CD4 help for B cell antigen presentation and other APC presentation which is a reason? Insert why not both.gif…Since we can't target antigen specific T cells in vivo for death using pentamers because of the low avidity and cross reactivity with other TCRs, Atara's solution of ex vivo expansion and re infusion likely is the only non myleoablative way of redistributing T cell populations. Combo of Atara method with Ocrevus should be a clinical trial.

  • "Overall reconstitution patterns appeared similar for participants who did and did not achieve remission through 60 months
    post-AHSCT. While a few dramatic deviations from the mean of the long-term remission group were observed for individuals
    who did not maintain remission, no consistent changes in cell
    subtypes studied characterized all or any of the three categories of
    disease reactivation. These changes could reflect true biology or
    could be spurious findings unrelated to disease activity or adverse
    events. Two of the seven participants were in remission at the time
    of this analysis and relapsed by MRI activity later on; therefore,
    it is possible that relevant biomarkers had not yet re-emerged."


    Ps: One year ago today i was taking citoxan and atg for Hsct

  • Cyclophosphamide was used against Hodgkin's Lymphoma, until less damaging treatments evolved. Hodgkin's Lymphoma is a B cell cancer, some versions being Epstein Barr infected immortal memory cells.

    • Well…Glad you ask

      08-01-2018 cd19+ ….95 cell/mcL (1,5months post transplat)
      cd27+ ….6 cell/mcL

      28/02/2018 cd19+…..269 cell/mcL (3 months pos tranplant)
      cd27+…..3 cell/mcL

      09/10/2018 cd19+…..495 cell/mcL (almost 11 months post trans.)
      cd27+…..16 cell/mcL

      CD27+ 1,4% as of total lymphocytes

      I was freaking out because cd27+ had risen more than 5 times

      I talk to neuro about that and the criteria for doing NMO 0,1%

      cd27+, protocol but he said that they are different diseases (Ms

      and Nmo) and you should not aplly the same protocol in is words

      " In NMO its down, down ,down, with the cd27+"

      Them i remmenber to see this study which is a much more ablative

      Hsct and my mumbers are in line with the b cell compartment


      Next tuesday i am gonna do full panel of CBC, T cells and b cell

      Exactly 1 year post tranplant i will let you know

      In the mean time i will be vigilant (last mri 2 months ago was

      Obrigado por tudo

    • Yes I noticed that the memory cells were quick to repopulate in the Storek paper. In NMO they were retreating at about 8 monts based on the repopulation kinetic and it is is a different treatment to rituximab on which the Kim et al papers are done

      Also we have no evidence that blood memory cell levels are important in MS.

      In some people there is a memory cell expansion due to EBV. I dont know if this influences activity.

    • In Hsct Ebv induced cd27+ expassion is mainly cause by t cell depleting agents ATG, Alemtuzumab

      I was doing weekly Ebv viral loads in IPO câncer center in Lisbon and the head heamatologish was surprise to see Ebv loads always bellow the detected level

      He said thats not usual When you use ATG


    • Please see my post ATG is B cell depleting (although more T cell targeting) and alemtuzumab is certainly B cell depleting.

      I also forgot to add, if you have ablative HSCT you have to regenerate you childhood immunities against common viruses and therefore you will expand up memory T and B cell subsets as your new immune system regenerates.

  • New (16/11): Immune reconstitution therapy (IRT) in multiple sclerosis: the rationale

    IRT modalities were shown to induce long-term remission of MS that, in some cases, is close to the definition of a "cure." There are cohorts of patients having been treated with the IRTs, alemtuzumab, and HSCT, who experience-under these modalities-no evidence of disease activity (NEDA) for over 10 years. Most importantly, IRTs cause radical changes in the lymphocyte repertoire after the reconstitution phase that may explain the long-term beneficial effects of IRT and the possibility of re-induction of self-tolerance to self/myelin antigens. In comparison, a chronic treatment cannot result in cure of the autoimmune reactivity, because it only blocks the immune system, as long as it is given; it cannot therefore radically affect the immunopathogenesis of the disease. The risks of adverse events related to immune suppression (such as opportunistic infections and secondary malignancies) with IRTs are lower and front-loaded, whereas the common side effects of chronic immunomodulation are higher and accumulate with time. In conclusion, IRT provides a novel concept for MS therapy with substantial advantages over chronic immunosuppression. IRT therapies have shown a significantly higher level of efficacy in MS. The "Holy grail" of the treatment of autoimmunity, which is to re-induce the disrupted self-tolerance, seems to be achievable-at least in part-with this approach. Moreover, the benefits of IRT, administered in short pulses, include significantly higher adherence to treatment and lower risks for accumulative side effects that are typically associated with chronic immunosuppression.

    • Thanks you have summarised the paper.

      I think they could further and I think that the lymphocyte repopulation is sterotyped, T cells repopulate in one way and B cells repopulate in another. We have just finished a review on cladribine and how it works as a SIRT (a selective IRT). Obviously it does not have the history of alemtuzumab or HSCT so we are hypothesizing the reality will transpire over the next few years

  • Hi,
    So I’m in relapse, 5 years post AHSCT. I had CTX-ATG. Should I check current levels of CD19+ and CD27+? How about EBV and Ch.P? I’m looking for a follow up treatment. Considering Ocrelizumab / Rituximab, Alemtuzumab or…. Tecfidera?
    Dziekuje 🙂

    • Reading further I can assume that CTX and ATG has depleted both T and B cells, therefore I was in remission for over 4 years. I have just checked my EBV and I have Igg 171 U/ml ang Igm <10 U/ml. So considering if I caught EBV again, after HSCT.
      To summarise I think that in my case Cladribine or Alem could do the job (depleting T and B). Any thoughts?

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