CSF – Prof G's MS Blog Archive

Clash of the Titans: the HILO study

Barts-MS rose-tinted-odometer: ★★★ (I am seeing blue and Spanish yellow today)
Roche blue (#0066CC) & Novartis Spanish Yellow (#F7B516)

Yes, I really do think that intrathecal (within the meninges that cover the brain and spinal cord) or CNS resident B-cells and plasma cells are pathogenic in MS. In other words, the cytokine or chemicals B-cells and plasma cells produce, in particular their antibodies, are what is driving some of the pathologies of smouldering MS. The evidence to support this hypothesis is well rehearsed on this blog and is the reason why we are testing high-dose ocrelizumab (more CNS penetrant) vs. standard-dose ocrelizumab (less CNS penetrant) against each other in two head-2-head studies. It is also the reason we are testing cladribine’s (CLADRIPLAS and CLAD-B) and ixazomib’s (SIZOMUS) effects in intrathecal B and plasma cell markers. Yes, I really do think we need to scrub the CNS clean of B-cells, plasma cells and their products, in particular the oligoclonal IgG bands.

I am therefore proposing a new study; the HIgh-dose versus LOw-dose anti-CD20 study or HILO Study.

In this study, I propose testing high-dose or double-dose ocrelizumab vs. standard or intermediate-dose ocrelizumab vs. low-dose ofatumumab against each other over two years and measure their impact on end-organ damage markers (slowly expanding lesions and brain volume loss) and on CSF markers of B-cell, plasma cell and microglial activity. The latter will include free kappa and lambda immunoglobulin light chains, OCBs, soluble CD14, etc. This will answer at least from a biomarker question whether or not we need CNS penetration of anti-CD20 monoclonal antibodies to target this component of smouldering MS. The following would also answer the question of whether or not you as a person with MS would want to be treated with high-dose or low-dose anti-CD20 therapy?

Would you want to be randomised into this study?

This study would be a clash of the titans; Roche vs. Novartis. Who would win? It really is not that important as Novartis is a major shareholder in Roche and hence when Roche makes a profit so does Novartis. The real winners will be people with MS, the data will allow them to make an informed decision about whether or not they want to go beyond NEIDA (no evident inflammatory disease activity) and be on a treatment that tackles the smouldering B-cell and plasma-cell driven processes within their brains and spinal cords.

SHOULD WE DO THE HILO STUDY?

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General Disclaimer: Please note that the opinions expressed here are those of Professor Giovannoni and do not necessarily reflect the positions of the Barts and The London School of Medicine and Dentistry nor Barts Health NHS Trust and are not meant to be interpreted as personal clinical advice.

St Elsewhere

Barts-MS rose-tinted-odometer: zero ★’s

St Elsewhere is a euphemism in medicine for a fuck-up done under someone else’s watch, i.e. at St Elsewhere’s hospital. In this week’s NEJM there is a tragic case of a 22-year old Pakistani man who was misdiagnosed as having MS at St Elsewhere, started on natalizumab, which was ineffective, before being switched to interferon beta. By the time he was admitted to UCSF he was in extremis. A relatively standard workup revealed this gentleman had a tumour; a CNS germinoma that had spread to the meninges or coverings of the brain and spinal cord. On reviewing his initial presentation there were so many ‘red flags’ it is hard to understand why he was misdiagnosed. Tragically this poor man now has ‘substantial neurologic disability, for which he received assistance with all activities of daily living’. This case is a tragedy because CNS germinomas when caught early and treated have a reasonable prognosis with a 10-year survival rate of about 70%.

Minter et al. Stalking the Diagnosis. N Engl J Med. 2021 Apr 1;384(13):1262-1267.

If only this patient had had a lumbar puncture and CSF analysis done as part of his initial diagnostic work-up the correct diagnosis would have been made. The question arises whether or not this gentleman had medical insurance or not? He was a delivery driver and had only recently immigrated to the USA. To be fair to the neurological team looking after him a diagnostic short-cut may have had to be made because he simply didn’t have the financial resources to pay for the diagnostic tests. I suspect the latter is likely to have happened and maybe the title of this post should have been St Poor.

I want to remind you that if you have been diagnosed with MS you may not have MS. In the study below approximately 1 in 5 people diagnosed with MS don’t have MS. This figure is much higher than previous studies. I have always quoted the Danish post-mortem studies that suggest that about 1 in 20 patients are misdiagnosed. Maybe Danish neurologists are simply better at diagnosing MS compared to their American colleagues?

There is no one test that can be done to diagnose MS. MS is diagnosed by combing a set of clinical and MRI findings, electric or neurophysiological investigations and laboratory tests. If these tests fulfil a set of so-called MS diagnostic criteria the Healthcare professional (HCP) or neurologist makes a diagnosis of MS.

The underlying principle of making a diagnosis of MS is showing dissemination of lesions in space and time and excluding other possible diagnoses that can mimic MS. The diagnostic criteria have evolved over time from being based purely on clinical attacks to those including electrical and spinal fluid tests to the modern era in which we use MRI to help confirm dissemination in time and space.

Dissemination in time means at least two attacks or two MS lesions occurring at least 30 days apart.

Dissemination in space means lesions occurring in different locations, for example, the optic nerve and spinal cord.

The electrical or neurophysiological tests are called evoked potential (EPs) and test electrical conduction in a particular neuronal pathway. They can be useful to show the effects of lesions in pathways that are not evident on neurological examination or seen on MRI. The EPs can also show slow electrical conduction which is one of the hallmarks of diseases that affect myelin, the insulation of nerves that are responsible for speeding up electrical conduction.

The laboratory tests are typically done to exclude other diseases that can mimic MS. One test that is useful in helping to make the diagnosis of MS is examining the spinal fluid for the presence of oligoclonal immunoglobulin G or IgG bands (OCBs), which are the fingerprint of a specific type of immune activation within the central nervous system (CNS). The OCB fingerprint is relatively specific for the diagnosis of MS in the correct clinical context. Please note OCBs can are found in infections of the nervous system and other autoimmune diseases, therefore, the presence of OCBs are not diagnostic on their own.

When CSF is sent to the laboratory they also measure the protein, glucose, lactose and do a cell count. An often the spins the cells out of the CSF and examine them to make sure they are abnormal. I suspect if this patient had had a CSF examination it may have been abnormal, which would have led the clinicians to the correct diagnosis, which will have allowed him to be treated differently; importantly, treated early and he may not have become profoundly disabled.

Why is getting the correct diagnosis of MS so important? Firstly, some of the treatments for MS have life-threatening complications; you don’t want to expose people without MS to these complications. Some diseases that mimic MS can be made worse by MS DMTs. This latter is particularly relevant for NMO or neuromyelitis optic. Patients with NMO misdiagnosed as having MS get worse on many of the MS DMTs. Finally, a diagnosis of MS has many psychological, social, financial and economic implications for people. Just having a diagnosis of MS, even if you turn out to have benign MS in the future, has implications for the person concerned. For example, it may affect your life choices and may impact your ability to get insurance cover to name to obvious examples. I would, therefore, advise you to make sure you have MS and not an MS mimic.

The most common MS mimics:

The evolving definition of MS based on diagnostic criteria:

Clinical criteria only:

Schumacher, et al. Problems of Experimental Trials of Therapy in Multiple Sclerosis: Report by the Panel on the Evaluation of Experimental Trials of Therapy in Multiple Sclerosis. Ann N Y Acad Sci 1965;122:552-68.

Clinical, EPs and CSF analysis:

Poser, et al. New diagnostic criteria for multiple sclerosis: guidelines for research protocols. Ann Neurol 1983;13:227-31.

Clinical, EPs, CSF analysis and MRI:

Kaisey et al. Incidence of multiple sclerosis misdiagnosis in referrals to two academic centers. Mult Scler Relat Disord. 2019 May;30:51-56.

BACKGROUND: Multiple Sclerosis (MS) specialists routinely evaluate misdiagnosed patients, or patients incorrectly assigned a diagnosis of MS. Misdiagnosis has significant implications for patient morbidity and healthcare costs, yet its contemporary incidence is unknown. We examined the incidence of MS misdiagnosis in new patients referred to two academic MS referral centers, their most common alternate diagnoses, and factors associated with misdiagnosis.

METHODS: Demographic data, comorbidities, neurological examination findings, radiographic and laboratory results, a determination of 2010 McDonald Criteria fulfillment, and final diagnoses were collected from all new patient evaluations completed at the Cedars-Sinai Medical Center and the University of California, Los Angeles MS clinics over twelve months.

RESULTS: Of the 241 new patients referred with an established diagnosis of MS, 17% at Cedars-Sinai and 19% at UCLA were identified as having been misdiagnosed. The most common alternative diagnoses were migraine (16%), radiologically isolated syndrome (9%), spondylopathy (7%), and neuropathy (7%). Clinical syndromes and radiographic findings atypical for MS were both associated with misdiagnosis. The misdiagnosed group received approximately 110 patient-years of unnecessary MS disease-modifying therapy.

CONCLUSION: MS misdiagnosis is common; in our combined cohort, almost 1 in 5 patients who carried an established diagnosis of MS did not fulfil contemporary McDonald Criteria and had a more likely alternate diagnosis.

CoI: multiple

Twitter: @gavinGiovannoni Medium: @gavin_24211

CSF vs. Blood neurofilaments

This blog post explains why blood neurofilament level monitoring is not ready for primetime and may never quite replace CSF NFL monitoring in pwMS.

Although blood neurofilament light chain (NFL) levels are a promising biomarker for monitoring MS inflammatory disease activity there are issues that need to be resolved.

We and others have shown that blood levels correlate with spinal fluid levels with a correlation coefficient of ~0.7. This translates to an R-squared value of ~0.5 and tells us that only 50% of the blood levels can be explained by what is detected in the lumbar CSF. We have many examples of patients low NFL levels in the CSF with high blood levels and the converse, i.e. high CSF levels and low blood levels. I think the discrepancy in these individual cases may indicate real biology.

CSF is an admixture of fluids derived from the (1) choroid plexus, where most of the CSF is made, (2) the so-called interstitium or substance of the brain and spinal cord, (3) the meninges and (4) the blood vessels that traverse the subarachnoid or CSF space.

The typical volume of CSF in adults is 125–150 ml. This turns over approximately four times a day, i.e. the daily volume of CSF produced is approximately 600 ml. CSF production and flow are subject to diurnal variation; flow rates of ventricular fluid can vary by as much as a factor of 3.5, with minimum CSF production occurring around 1800h (12±7 ml/h) and a maximum at approximately 0200h (42±2 ml/h).

The integrity of these barriers and the flow of CSF determine the content of CSF protein. The CSF constituents are not uniform and depend on the site from which the CSF is sampled. There is a head-to-tail concentration gradient for total protein along the axis of the CNS, with the lowest concentrations in ventricular fluid and the highest concentrations in the fluid from the lumbar-sacral sac from where the CSF is sampled during a lumbar puncture.

Physical activity influences the CSF protein concentration; subjects who have been lying down for prolonged periods have higher CSF protein concentrations than active subjects. It is important to consider this when interpreting the results of CSF analyses from subjects who have been on their backs for more than 24 hours.

The lumbar subarachnoid space is a cul-de-sac. CSF obtained from lumbar punctures does not necessarily provide accurate information on inflammatory or pathological events in the brain that occur beyond the CSF outflow pathways of the fourth ventricle, i.e. events in relation to the surface of the brainstem, cranial nerves and cerebral hemispheres. In addition, the extracellular space of lesions deep in the brain may not necessarily communicate with all parts of the free CSF space.

For these reasons, spinal NFL levels are more likely to represent spinal cord disease and not brain pathology. In comparison, blood levels are more likely to integrate what is being produced from the whole CNS and the peripheral nervous system. Yes, NFL is not specific to the CNS. Therefore NFL from peripheral nerve disease will affect blood levels.

Another issue is that some pwMS mount an autoantibody response against NFL proteins. These antibodies may reduce the circulating NFL levels and hence may affect peripheral blood levels by artificially lowering NFL levels. The anti-NFL antibodies tend to occur in people with more advanced or progressive MS and hence may explain why peripheral blood NFL levels are so low in people with progressive MS.

Another factor is the circulating blood volume; the study below shows the larger your blood volume the lower your peripheral blood NFL levels will be. Height and body size will need to be taken into account when interpreting blood levels.

A critical factor that needs to be determined is how long does NFL circulate in the periphery and what is the mechanism of its clearance and breakdown. Without knowing the latter we will continue to have difficulty interpreting what peripheral blood NFL levels mean.

Could peripheral blood levels increase as part of normal biology? I was recently told at a meeting that peripheral blood NFL levels increase as a result of running a marathon (unpublished data). Why? Could this be due to microtrauma to peripheral nerves or the neuromuscular junction? Or due to increased release from the brain? We know that minor head injuries raise NFL levels. Could the excessive bobbing around of the brain in the skull during marathon running result in a small NFL leak? Marathon running also affects peripheral fluid balance; the increased NFL levels could be related to an alteration in blood volume or the breakdown and clearance of the NFL in the periphery.

So switching from CSF to peripheral blood monitoring of NFL levels is not going to happen until we have answered some of these questions. In addition, it is unlikely that we are going to make treatment decisions based on isolated peripheral blood NFL measurements. NFL levels will be one of many factors that will need to be taken into account when making clinical decisions. Serial or longitudinal NFL measurements with an area under the curve analysis will be more valuable than a single peripheral blood measurement.

These are some of the reasons why peripheral blood NFL levels are not quite ready for primetime and clinical practice. Do you agree? This is why we at Barts-MS will continue to do lumbar punctures and CSF measurements of NFL in our patients for the foreseeable future. So if you come to see us in our clinic we may offer you a monitoring lumbar puncture.

In addition, to NFL levels there are many other biomarkers that can be measured in CSF that will provide us with information on smouldering MS disease activity. This is why Barts-MS is ideally positioned to tackle the challenge of defining and studying smouldering MS.

Manouchehrinia et al. Confounding effect of blood volume and body mass index on blood neurofilament light chain levels. Ann Clin Transl Neurol. 2020 Jan 1. doi: 10.1002/acn3.50972.

Blood Neurofilament light chain (NfL) has been suggested as a promising biomarker in several neurological conditions. Since blood NfL is the consequence of leaked NfL from the cerebrospinal fluid, differences in individuals’ Body Mass Index (BMI) or blood volume (BV) might affect its correlation to other biomarkers and disease outcomes. Here, we investigated the correlation between plasma NfL, BMI, and BV in 662 controls and 2,586 multiple sclerosis cases. We found a significant negative correlation between plasma NfL, BMI/BV in both groups. Our results highlight the potential confounding effect of BMI/BV on associations between blood NfL and disease outcomes.

CoI: multiple

Beyond the B-cell

Do we have the right cell target in MS? Yes and no; we need a multicellular approach.

Recently the attention in MS has been on the B-cell as if it was the holy grail of MS treatments. It is not.

In several posts, over the last few weeks, I have made the case that the B-cell is important, probably as an antigen presenting cell, but it is not the ‘be all and end all’ of MS treatments. It is clear that rebound post-natalizumab is driven my B-cells and the positive data on the first BTK inhibitor would indicate that the B-cells are working via the B-cell receptor on antigen presentation. If only we knew what these antigens were we would have a much better handle on the cause of MS.

I know this science stuff is hard, but it is important. At the end of the day, the nut and bolts of MS must be molecular; molecules mean treatment targets and potentially more focused and hopefully better and safer treatments in the future.

I have stressed that simply targeting B-cells in both the periphery and central nervous system will not be enough to effectively treat MS in the long-term. When we look at end-organ damage markers in pwMS who are on B-cell therapies they have ongoing brain volume loss, albeit at a lower rate, and enlarging lesions (T1 black holes), which are both indicative of ongoing smouldering MS. So what do we need to do? I have provided circumstantial evidence that NIRTs (non-selective immune reconstitution therapies) have a slight edge on the B-cell therapies and this may be because they are also targeting T-cells. The latter, however, comes at a price of greater adverse events in relation to immunosuppression. The proportion of MSers on NIRTs who experience disability improvement seems higher when compared to the anti-B cell agents, which indicates that NIRTs are doing something else over and above their effect on the B-cell compartment. However, based on their overall safety profile it is unlikely that the NIRTs (alemtuzumab & HSCT) will be a therapeutic strategy that the wider MS community will adopt with vigour. Although from comments on this blog there is an informed group of MSers who feel hard done by because their HCPs won’t offer them the option of using NIRTs first-line, i.e. very early in the course of their disease when they have the most to gain from these therapies.

Is there anything else we can do to improve on the profile of B-cell therapies to make them better? Yes, I think there is. Targeting the plasma cell,in addition to the B-cell. Data on plasma cells goes back decades and surprisingly the plasma cell has never been a major therapeutic target in MS. John Prineas, one of my MS heroes, has always stressed the importance of the plasma cell in MS. His paper below from 1978 documents just how enriched the brains of MSers are with this population of cells. What is often not stressed is that the biology of plasma cells is so so different to the B-cell, which opens up new therapeutic targets that are quite different to those in the T and B cell compartments. More on this topic another time.

You are aware of the recent publication showing that about 55% of Polish MSers treated with intravenous cladribine lost their oligoclonal bands 10 or more years after treatment and if they did lose their OCBs they tended to have lower EDSS scores. We have known for years that MSers, with either relapse-onset or primary progressive diseases, who don’t have OCBs do better. There is also evidence from biomarker and pathology studies that the OCBs may be driving several of the disease processes that have been linked to advanced or progressive MS, i.e. microglial activation and grey matter pathology. Based on these observations, we hypothesise that OCBs are very likely to be pathogenic in MS, which is why we are embarking a research programme to try and target the plasma cells within the CNS of MSers. Do you think we are crazy?

To get a handle on the plasma cell we are going to have to study what happens in the spinal fluid. There are simply too many plasma cells in the periphery which will drown out any signal from the CNS. To participate in the studies we are planning we will have to perform serial, annual, lumbar punctures or spinal taps to see if our add-on therapy is killing and/or reducing the number of plasma cells in your brain and spinal cords. The good news is that we have de-risked the lumbar puncture with the use of atraumatic needles and screening. I never thought I would be saying this but most of our patients don’t mind having LPs, particularly when they understand the reason behind the LP. CSF neurofilament levels are now part of our prognostic profile of MSers at baseline and we are increasingly using them to assess response, or lack of response, to treatment. So if you want to be treated and treated-2-target beyond NEDA, and beyond the B-cell, then having an LP is important.

We hope our proposed plasma cells studies will lead to a mindset that goes beyond the B-cell to target some of the mechanisms that are responsible for smouldering MS.

Prineas & Wright. Macrophages, lymphocytes, and plasma cells in the perivascular compartment in chronic multiple sclerosis. Lab Invest. 1978 Apr;38(4):409-21.

Perivascular cells in CNS tissue from six multiple sclerosis (MS) patients and a patient with motor neuron disease were examined by light and electron microscopy. Lymph node tissue from one MS patient was also examined. CNS perivascular macrophages in both MA and motor neuron disease were found to closely resemble free macrophages elsewhere in the body except that they often contained unusually large primary lysosomes. Cytoplasmic inclusions consisting of membrane-bound stacks of curved linear profiles, presumed to be a product of myelin degradation, were constantly observed in microglia in MS plaques but were rarely observed in perivascular macrophages in the same area. Unidentified cylindrical bodies were observed within cysternae of rough endoplasmic reticulum in some lymph node cells. Quantitative studies of the perivascular cell population in one MS case revealed, in histologically normal white matter 260 lymphocytes and 178 plasma cells per cubic millimeter of fresh tissue. Typical chronic plaque tissue without obvious inflammatory cell cuffing contained 1772 plasma cells per cubic millimeter of fresh tissue. No plasma cells were observed in the CNS in motor neuron disease. The results of this study suggest that perivascular macrophages in the CNS represent a specialized population of monocyte-derived free macrophages, that these cells differ functionally from microglial cells, and that the digestion of myelin breakdown products in MS requires the participation of both cell types. The results also suggest that in some chronic MS cases there is a large, permanent population of CNS plasma cells that persists, like the elevated cerebrospinal fluid IgG level in this disease, for the life of the patient, that these cells, rather than inflammatory cells in fresh lesions, are the major source of this raised IgG, and that the existence of such a population of cells may indicate the continuing expression of antigens in chronic MS lesions in the absence of fresh lesion formation.