Natalizumab biopsies, is the T cell hypothesis all wrong?


Natalizumab, a recombinant humanized monoclonal antibody directed against the α4 subunit of the integrins α4ß1 and α4ß7, has been approved for the treatment of active relapsing-remitting MS. It has been suggested to work by blocking T and B cells and perhaps monocyte entry into the central nervous system. This inhibits relapsing disease but the side effect seems to expose people to the risk of progressive multi-focal encpehalopathy. If this is true one may guess that if you looked at a MS brain from someone treated with natalizumab, that it would be full of remyelination as the disease is stopped and repair could occur. However the repairers have challenged the view that the pathologists can easily pick up repair as they have suggested that it can be invisible and not be associated with thinly remyelinated axons. So the question arises what happens when you do biopsies in natalizumab treated people with MS?

It seems like it does nothing according to the Pathologists. So how do we explain the activity?

CNS inflammation after natalizumab therapy for multiple sclerosis: A retrospective histopathological and CSF cohort study. Brain Pathol. 2021 May 6:e12969. doi: 10.1111/bpa.12969. 

Natalizumab, a recombinant humanized monoclonal antibody directed against the α4 subunit of the integrins α4ß1 and α4ß7, has been approved for the treatment of active relapsing-remitting MS. Although natalizumab is a highly beneficial drug that effectively reduces the risk of sustained disability progression and the rate of clinical relapses, some patients do not respond to it, and some are at higher risk of developing progressive multifocal leukoencephalopathy (PML). The histopathological effects after natalizumab therapy are still unknown. We, therefore, performed a detailed histological characterization of the CNS inflammatory cell infiltrate of 24 brain specimens from natalizumab treated patients, consisting of 20 biopsies and 4 autopsies and 21 MS controls. To complement the analysis, immune cells in blood and cerebrospinal fluid (CSF) of 30 natalizumab-treated patients and 42 MS controls were quantified by flow cytometry. Inflammatory infiltrates within lesions were mainly composed of T cells and macrophages, some B cells, plasma cells, and dendritic cells. There was no significant difference in the numbers of T cells or macrophages and microglial cells in lesions of natalizumab-treated patients as compared to controls. A shift towards cytotoxic T cells of a memory phenotype was observed in the CSF. Plasma cells were significantly increased in active demyelinating lesions of natalizumab-treated patients, but no correlation to clinical disability was observed. Dendritic cells within lesions were found to be reduced with longer ongoing therapy duration. Our findings suggest that natalizumab does not completely prevent immune cells from entering the CNS and is associated with an accumulation of plasma cells, the pathogenic and clinical significance of which is not known. As B cells are considered to serve as a reservoir of the JC virus, the observed plasma cell accumulation and reduction in dendritic cells in the CNS of natalizumab-treated patients may potentially play a role in PML development.

This study looks at brain tissue from people treated with natalizumab and seems to suggest that natalizumab does not do much in the brain and it does not stop T cells entering the brain. If really true then it questions the basis on how natalizumab is thought to work and in doing so could suggest that the underpinning animal work is not correct.

Dare I comment on this and perhaps in the new era I should perhaps not. However, it is an H bomb to the fundementals of MS drug biology.

They say “Of the 24 natalizumab‐treated patients, 13 responded to the therapy (labeled as resp.), while 4 patients did not (labeled as non‐resp.). Non‐response was defined as presenting with relapses and/or new lesion formation on magnetic resonance imaging (MRI) during treatment with natalizumab. In 7 patients, no information on therapy response was available. Controls (labeled as MS) included 21 MS patients with no prior natalizumab therapy, and a disease duration comparable to natalizumab treated patients (n = 11 biopsies and 10 autopsies). In the four autopsy cases, the cause of death was a fulminant MS disease course in two patients (treatment failure), cardiac infarction in one and recurrent infections in the other. Brain biopsies were taken for diagnostic reasons, e.g. to rule out progressive multifocal leukoencephalopathy (PML), a known severe side effect of natalizumab treatment, or due to clinical deterioration after stopping natalizumab therapy.” Tissue in all patients showed inflammatory demyelinating lesions consistent with MS. Furthermore, the interval between the last natalizumab infusion and biopsy/death varied between 20 days and more than 5 years. They did not find a difference in T cell numbers between those sampled at less than 14 weeks verses those at more than 14 week.

They say “Taken together, our results suggest that T cell infiltration into the brain parenchyma may occur despite natalizumab therapy”.

Playing devils advocate This is more information that MS has nothing to do with MS, and could imply it has nothing to do with controlling PML. However, I am not so sure.

But let’s start in the cerebrospinal fluid in the non-biopsied, living cohort and there we get what we expect….or do we?. There is a reduction in the number of T cells notably a reduction of CD4 T cells and no significant effect on CD8 T cells, but naive CD8 cells were reduced. Therefore evidence to suggest CD4 T cells are important. There are not enough people with high levels of non antibody-producing B cells (CD19+, CD138-) to make this significant. CD138+ antibody producing cells stay the same. They do not specificallly look at memory B cells…so we can’t make a call and so it does not destroy the hypothesis that memory B cells are important in MS.

However, it has to be said that this has been done before and the number of CD4, CD8 and CD19 B cells dropped (Stüve O et al. Ann Neurol. 2006;59:743, Kowarik MC et al. Neurotherapeutics. 2020. doi: 10.1007/s13311-020-00975-7, so it is not the same. In this study they look at proportions, the B cells drop and the memory CD8 T cells increase and the CD4 cells stay the same and others drop whilst others increase, but as we have said before if you are looking at proportions if one goes down the others go up and if one goes up the others do down and therefore it is difficult to make firm conclusions.

However, let’s go to the biopsies. You can make of this what you will, but one could argue that looking years after the sessation of natalizumab is not going to give you the answer and some people were examined years after natalizumab but others were not. However, I will simply say this. These people were biopsied and therefore one imagines there was something on an MRI scan to trigger this investigation in a certain place in the first place. However, that the people are having biopsies or were dying rather says, something was not working. Some of the biopsies were done for concern of PML, but PML cases were excluded from the study. So one suspects that cases with disease activity were selected and therefore you would expect to see inflammatory cells. The pictures show cells inside tissue. Here CD4 cells predominate, yet it has been argued by the pathologists that cells in tissue are CD8 T cells. So I will sit on my fence. There is clearly animal data in support of natalizumab inhibiting migration of cells into the CNS.

Disclaimer: please note that the opinions expressed here are those of the authors and do not reflect the positions of the Barts and the London School of Medicine and Dentistry, nor Barts Health NHS Trust or Queen Mary University London

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  • They say: “We consider the varying intervals between the last natalizumab infusion and biopsy/autopsy to be the most important limitation, yet our study contains to date the most comprehensive collection of CNS specimens obtained after natalizumab treatment.” I find this justification quite incredible given the mean(!) delay between Natalizumab cessation and biopsy was 185 days. I’m sorry to say this, and I’ve had excellent discussions in the past with most of the authors, however a gap of such duration with an agent that has a half-life of 16 (+/-4) days, and a well-known risk of rebound disease, significantly limits any conclusions drawn from this paper (did they inadvertently describe some elements of rebound activity here?).

      • The biopsies were done for reasons of clinical care and therefore we do not need to question the ethics. The pathology would have been done for this reasons too, but there was tissue left for research, which is an offshoot. What profK is saying by the time the study was done the brakes would have been removed.

        However, these people had been switched from natalizumab and no doubt for good reasons but as ProfK says by the time the biopsies were done the influence of natalizumab may have diminished and as there was disease activity.

  • This is gold thanks for this doc Mouse

    Prof k has a good point


    They say:

    Furthermore, the interval between the last
    natalizumab infusion and biopsy/death varied between
    20 days and more than 5 years.

    (so there where people in the 16 days(+-4) half life period there brain biopsy should be quite diferent from the others9

    Again they say

    After discontinuing natalizumab treatment, a reduction
    in free CD49d receptor binding sites on PBMCs
    was described for a further 3½ months as compared to

    levels before the first injection.(10) To determine whether
    natalizumab activity has an influence on CNS T cell
    inflammation, T cell numbers in active demyelinating biopsies
    until 3½ months after the last infusion were compared
    to T cell numbers with treatment discontinuation
    longer than 3½ months. No difference in T cell numbers
    was found when comparing these two groups (Figure 1B;
    interval between last natalizumab infusion and biopsy
    or autopsy 3½
    months median: 53.7 cells/mm2).

    To me this means that a treament efects is visible even after 3,5 months …However before and after 3,5months no efects on t cells

    In discussion part

    T cell numbers were
    not significantly reduced as compared to non-treated
    controls. Moreover, T cell numbers were also not lower

    when the interval between the last natalizumab infusion
    and the biopsy or autopsy was short, indicating
    that T cells may enter the CNS despite natalizumab

    So Prof K your reason does not fully hold

    • You cannot make a study where obviously treatment 1) did not work ( this is why a biopsy was take) because of anti-natalizumab antibodies or other factors (natalizumab does not have a 100% effect against relapses, there are patients that do not respond) or 2) where you have a rebound effect (long interval) and then say: look, this is how natalizumab works… if anything this paper supports the T cell hypothesis, i.e. you find T cells when you have inflammation.

      • You cannot make a study where obviously treatment 1) did not work

        So in your view you only can make astudy if and when patients respond 100%?

        20 days its a long interval?

        • If you want to see how medication works you have to study what happens when it works. If you have for example antinatalizumab antibodies, time since the dose not play any role, the medication does not work, you get new inflammation that you have to do a biopsy on to exclude pml and if its not pml, you report what you see when medication obviously does not work.
          If you want to test how natalizumab works, you have to include people who are under natalizumab and dont have new inflammation but difficult to have biopsies.
          Of course its a very interesting study but cannot say anything about the drug mechanism.

          • If you want to see how medication works you have to study what happens when it works.

            Understand relapse aftr cessation of treatments are surely informative

      • Good point, I agree but it was stated that many were responders and as for T cells you find B cells and lots of macrophages

        • I agree, I dont exclude b cells and monocytes, just say that t cells are there.
          Regarding responders, why did the three patients with inactivated biopsies receive corticosteroids and plex? Why did they take the biopsies in responders, was it contrast enhancement in old lesions; slowly expanding lesions mistaken for pml? More clinical data are needed.

    • The point is taken but rebound occurrs before 14 weeks in some people and therefore the level of CD49d blocakage has dropped enough to allow cells to enter the CNS. I think to suggest that events in the CNS can relate to something given 5 years ago, is a hard link to make



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