ANDREW MCKEON: So my name is Andrew McKeon, and I'm a director of the Neuroimmunology Laboratory at Mayo Clinic and also a consultant in the Department of Neurology in the Autoimmune Neurology Clinic. I'm going to present a summary of autoimmune central nervous system disorders and the approach we take to the evaluation and management of patients affected by these disorders at Mayo.

So autoimmune central nervous system disorders, these are neurological disorders caused by an aberrant immune response, so you might think of multiple sclerosis in that situation when I mentioned that. But unlike MS, there's a specific antigen that is affected and is the target of the immune response in these patients. These disorders may be triggered in the setting of cancer where there is a nervous system protein expressed in the cancer that results in an appropriate immune response against the cancer, but you get some aberrant immune response against that same protein in the nervous system.

There are also examples of parainfectious autoimmune neurological disorders such as in Guillan-Barre syndrome but also more recently with post-HSV NMDA receptor encephalitis, but the majority of patients we see with these disorders, the trigger is unknown. Many of these patients have autoimmunity in their background, but their precise trigger and why it occurs at any particular moment is, a lot of the time, unclear. But these disorders are unified by one or more antibody biomarkers in serum or spinal fluid.

So how do the patients present? Typically these patients present with subacute onset symptoms, meaning the symptoms that evolve over days to weeks. So not over seconds to minutes like in stroke, and not like insidiously progressive over months to years like you might see in Alzheimer's disease, but something in between. Sometimes these patients have very rapid progression though. In other situations, have a fluctuating course. And these disorders can affect any neurological domain and so really anywhere from cortex to C fibers.

And some of these disorders fall into a category of a classical phenotype such as limbic encephalitis, but then you may have atypical phenotypes where patients don't quite fit into that classical textbook pattern, such as GFAP autoimmune encephalitis. And then in some circumstances you can have patients with multifocal disorders where this is not a classical presentation for any disorder. And with that self-acute onset and with the combination of different levels of the nervous system involved, autoimmunity or perineoplasty disorders may come to mind. So an example of that would be a patient with sudden onset chorea, late life, no family history but also onset of neuropathy at that time.

Sometimes the levels of the system and the associated disorders, we're referring to encephalopathy at the cortical level but also epilepsy in some patients with more of a rapidly progressive dementia phenotype, optic neuritis, retinopathy, basal ganglia disorders such as movement disorders, ataxias. And at the brainstem level, then diplopia and ataxia, at the spinal cord level, then myelopathies or myelitis, and then there can be some patients with myeloneuropathies.

Sometimes paraneoplastic disorders can present with lumbosacral radiculopathies, neuropathies. Classic example is a sensory neuronopathy that we see in paraneoplastic disorders with small cell lung carcinoma or sometimes gynecologic cancers. We don't see patients with ALS-type presentation. That tends to be exclusively a degenerative disorder, though sometimes patients are referred because they have kind of an encephalomyelopathy of unknown cause, and people are thinking anterior horn-style disease is on the list. So sometimes it's a disorder that you have to differentiate from ALS, and then neuropathies, myopathies, and then neuromuscular junction disorders such as myasthenia gravis and Lambert-Eaton syndrome.

So why do these disorders occur? Well, the examples are mentioned in one of my previous slides, but just to delve into a little bit of detail here as far as prototypic disorder of cancer related to neurological autoimmunity, we can glean a little bit about the pathophysiology and also a little bit about how these antibodies serve as biomarkers, and also how they may actually be disease causing themselves. So these IgG antibodies are generated in the setting of a T-cell-mediated disorder, then you can have development of cytotoxic T cells, which you also may have development of B cells and then B-cell maturation after stimulation by the T-cell arm.

And then you can have a variety of antibodies with different effects, but on the green side over here, this really emphasizes proteins that are the antigens that are intracellular. So they're in the nucleus cytoplasm or sometimes the nucleolus, and the protein is degraded in the proteasome. And then you get these polypeptides that are expressed on MHC class I on the cell surface, and then these are recognized by cytotoxic T cells rather than antibodies themselves.

But in this whole milieu of this immune response, you still get antibody generation, so these antibodies are still there. There are still robust biomarkers of whatever this particular response is. So this protein here could be a polypeptide sequence from the Hu protein and ANNA-1 autoimmunity or, say, Ma2 as an example, but really, the main players here are the cytotoxic T cells. And we see that [INAUDIBLE]. So antibodies, while not disease-causing in this scenario, are still very good biomarkers in the lab.

On the other hand, you have examples here of autoantibodies and their potential pathogenic effects where you have a cell surface receptor. So not in the cytoplasm, not in the nucleus or in the nuleolus, but where you have at least part of the protein expressed on the cell surface and the three-dimensional structure of the protein, as it is in vivo, becomes important. And so these antibodies are not directed against linear epitopes like we saw over here before these three-dimensional epitopes. Examples of that include NMDA receptors, aquaporin 4, water channels.

And these antibodies, as well as being CO biomarkers in the lab, can also have a role in disease, and there's more and more data coming out on that for various diseases, including NMDA receptor encephalitis where down regulation of the receptor is important. But in the other scenario, you could also have antibody-activated complement and causing a lot of inflammation like in neuromyelitis optica where the water channel aquaporin 4 is targeted.

So that's just to give you a kind of a broad overview of the pathophysiology and the antibodies as kind of broad groups. But the disease categories to consider are as follows in these patients, encephalopathy, other typical limbic encephalitis or some other examples that we've come to, some patients with a kind of a more of a dementia phenotype that might mimic CJD, epilepsy where it's a limbic encephalitis-type presentation in some situations, but the person presents exclusively with seizures, and then you can have neuromyelitis optica spectrum disorders, these inflammatory disorders of the optic nerve and spinal cord that mimic multiple sclerosis, autoimmune movement disorders such as choreas, for example, associated with CRMP5, paraneoplastic myelopathies such as associated with, again, CRMP5 or ANNA-1/anti-Hu.

And then a variety of different neuropathies, either paraneoplastic or idiopathic. Some of which are responsive to immunotherapy, such as Caspr2 autoimmunity and others that are poorly responsive to immunotherapy such as ANNA-1 or anti-Hu. And then you have myasthenia gravis and Lambert-Eaton syndrome, which are the neuromuscular junction disorders, either postsynaptic or presynaptic, and then myopathy such as necrotizing autoimmune myopathies.

So the risk factors are sometimes are nonexisting, but there can be clues. For example, in GAD65, neurological autoimmunity such as stiff-person syndrome or ataxias, or NMO, in neuromyelitis optica, patients very commonly have a history of autoimmune disease already established. So asking about that, asking about a lot of autoimmunity in the family can be helpful as well. And then for paraneoplastic disorders, cancer history and smoking history can be important also.

For patients where the person comes down with a subacute onset neurological problem and the cause is not immediately apparent, it's also important to focus on symptoms such as preceding flu-like symptoms that might be a clue to a systemic infection that then results in a post-infectious inflammatory state. And in some situations, there are antibody biomarkers we find in that scenario.

So evaluating further-- so there's two parts to this. It's really important to document the neurological examination. Doing mental status testing, for example, neuropsychometric testing, brain imaging, electrophysiology, as appropriate. And all of these are really targeted towards the phenotype that's determined from the history and the exam. It's also important then to think about-- OK, what are kind of the basic kind of tests that we should be doing for that particular phenotype? So subacute onset neurological disorder could be resulting from things that are very straightforward like vitamin B12 deficiency or folate deficiency or some other metabolic problem.

And then moving on to autoimmune disease, we can think about clues to autoimmunity such as thyroid antibodies, connective tissue disease antibodies. And then more [INAUDIBLE] are [INAUDIBLE] testing and neural antibodies in the serum or CSF. And then these other markers, proteins, cell count, IgG index and synthesize rate, and oligoclonal bands can be very useful as well. So these are typically-- we think about these things in multiple sclerosis, but these can also be very helpful for other inflammatory disorders of the nervous system, such as autoimmune or paraneoplastic disorders.

So thinking about the autoantibodies themselves in these two kind of broad groups, what are we talking about? Well, these green ones, where I was talking about earlier. Where you have this cytotoxic T-cell-predominant response, a lot of these fall into the groupings of antibodies that were originally described, these classical paraneoplastic antibodies. Small cell carcinoma is a big theme here because small cell neoplasms essentially express a lot of neural antigens, and sometimes you can have a multitude of these antibodies positive in a patient with underlying small cell.

You can see that there are some exceptions to this, including, particularly, for anti-Yo or PCA-1, where its a many cancers that affect women. And then I've highlighted kind of a few other ones here, which were ones that have kind of, generally, a pretty low level of cancer association but still kind of fall into this bracket of an antibody that's directed at us in intracellular protein where cytotoxic T cells and cytokines and so forth are likely to have a big role in pathogenesis. But the other ones that I haven't highlighted in that way, typically have kind of a 70% to 80% positive predictive value for cancer.

In contrast to a lot of this, synaptic autoantibodies have a lower cancer association such as Lgi1/CASPR2. We do encounter some occasional cases of thymoma and other cancer ties but probably less than 10% overall and [INAUDIBLE] for things like glycine receptor antibody and DPPX. The ones I've highlighted in blue here are kind of the opposite of what I just showed you on the other side. These actually have a stronger cancer association than the others, the NMDA receptors, AMPA receptors, GABA-B in particular.

So how are the antibodies detected then in the lab? So you've kind of determined your phenotype. You've kind of determined the type of evaluation I might be thinking about here as far as an antibody profile for a particular disease state such as encephalitis. So when the serum or spinal fluid comes into the laboratory, then we have to determine what type of testing to do. And a lot of the different antibody analytes that we've, either, discovered or brought in from labs elsewhere, we've really just evaluated a lot of different platforms to see which is the most often way to test for a particular antibody.

But, in general terms, a lot of these plasma membrane protein expressed at confirmation of epitopes, these 3D epitopes we talked about earlier, a lot of them are detected by cell-based assays, either observer-based here or by flow cytometry, either as a screening test or as a confirmation test. And then the real workhorse is this tissue-based immunofluorescence assay that you can see here at the bottom. And so this is [INAUDIBLE] brain tissue that's stained with the patient's serum or spinal fluid. And then you have a secondary antihuman antibody that labels with a fluoroform, and you've got these different patterns of staining of tissue. And so this is a very characteristic staining for PCA1 or anti-Yo that I mentioned earlier.

And then there are other methods for detecting autoantibodies in the lab. Some have some pitfalls-- so Elisa, radioimmunoprecipitation assays. So in a particular context, these can be helpful but they sometimes can be prone to false positives, particularly a kind of low antibody values. So examples of that will be say anti-CagA antibody, which we detect by radioimmunoprecipitation assay. We can detect this and up to about 2% of the general normal population, but at a very high value in a patient with Lambert-Eaton syndrome, this is generally indicative of underlying small cell lung cancer, so some assays require more clinical interpretation than others.

And then an example of how we might detect an autoantibody in the cell-based assay in an automated format in the laboratory is by flow cytometry. So typically flow cytometry is used or has been used for detecting antigens. So most people would be familiar with-- you give somebody rituximab, give a patient rituximab, and then you want to follow that CD19 or CD20 count expressed on B lymphocytes. And so the way that's done is the analyte that you're interested in is actually the antigen on the cell surface of the B cells. Is CD19 or CD20 present? And the detection antibody is an anticommercial anti-CD19 or anti-CD20 antibody.

In this scenario, we're doing it the other way around. So it's actually the antibody that is what the analyte of interest that we're trying to detect while the reagent that we're using to detect that is a cell that expresses either aquaporin 4 or MOG protein. And so that can be done by doing the following. You have the cell population that's transfected with the protein of interest, and then this is labeled with the patient serum or spinal fluid. And then you have a secondary antibody, [INAUDIBLE], so it gives off a signal.

And you can tell by looking at the different cell populations then what you're detecting here, and so, in this example here, you're looking at a negative spasm, where you're not getting this antihuman IgG signal at all. Whereas in this one here you're seeing, OK, I've got some population that don't express the GFP-type protein at all, and then I have this other population that do express it, but this population that does express it is also giving me this strong signal for the antihuman secondary. And then you can express that as a fraction and give yourself an index. So the reason I'm pointing this out is this is going to be sort of a unique and novel way of using flow cytometry for detecting autoantibodies and allows a kind of a high throughput format and rapid turnaround time for clinical practice.

So an example of how this kind of intersection between the clinical space and the laboratory space at Mayo has really worked in a disease state is neuromyelitis optica. So these were kind of the patients, the MS patients, that people really had a lot of difficulty managing because, really, they didn't respond to any classical MS drugs. These patients with NMO would have attack after attack of inflammatory neuritis, optic neuritis, or myelitis, and when that would end up with, very often, with blindness and being wheelchair-dependent no matter what people did to treat them.

But then through some studies by Dr. Weinshenker and Dr. Wingerchuk at Mayo, these were clinically classified as distinct from multiple sclerosis. And a lot of early clues were there, including the type of lesions that we're seeing on MRI imaging and also the autoantibody associations, and that these patients had a lot of thyroid autoimmunity and connective tissue diseases like lupus. And Dr. Lucchinetti observed that the pathology in these patients was different from multiple sclerosis, and that there was evidence of complement cascade activation and the lytic part of the complement cascade, C9, deposited around blood vessels, which was unique as compared to multiple sclerosis.

And then Dr. Lennon subsequently discovered in these original patients that have been described, this particular autoantibody pattern that when use neuromyelitis optica IgG, and then subsequently discovered that this was aquaporin 4 IgG or an antibody against the aquaporin 4 water channel. And then Dr. Lennon also subsequently showed that the antibody had functional effects, including activation of the complement cascade. And so then it was known that there were complement inhibitors, cascade inhibitors already out there for use in paroxysmal nocturnal hemoglobinuria.

So Dr. Pittock and Dr. Wingerchuk left this open label study in 2012 demonstrating a very promising effect from this. And then this was replicated in a phase three randomized control trial in 2019 that was published in the New England Journal of Medicine, and, subsequently, this drug became FDA-approved. And so like over the course of, essentially, 20 years you went from orphan disease to a drug that could potentially cure patients.

So that's what we're really aiming to do in our clinical practice as it intersects from the face-to-face practice on one side of 2nd Street in Rochester and in the laboratory setting across the street. So one of the other observations has been over time now is these biomarkers that I showed you earlier. As these have begun to accumulate in number, there's been an increase in recognition of these disorders, of autoimmune encephalitis in particular. And when you take a population of patients, in an epidemiologic sense, some decades ago, things would have broadly broken down into infectious encephalitis accounting for the majority of cases, autoimmune encephalitis accounting for minority, and then also having this large group of patients with an undiagnosed or encephalitis of unknown cause.

But this has changed over time, and, essentially, now infectious encephalitis, autoimmune encephalitis of a pretty similar incidents and prevalence, and the number of unknown cases is dropping. But this is really, we think, a reflection of case recognition, and a lot of that probably has to do with increasing awareness because of a lot of publications around this but also diagnostic antibody markers in the laboratory.

And so just to make a couple of points for clinical practice about these autoantibodies and their utilization. And so this is an example of a patient with limbic encephalitis, this coronal flair MRI. And you can see that T2 hyperintensities in the limbic regions, and you can see the types of symptoms that these patients have. But one principle that is important for these patients is a diagnosis that occurs quickly, so that's both clinical and radiological recognition but also then serological confirmation. And we find that a profile-based approach is most helpful here, taking all of the autoantibodies that are pertinent to this particular presentation, and testing for these at the same time.

Why? Because time is brain in terms of getting patients on a track for treatment as quickly as possible, being able to counsel patients as quickly as possible as to the likely outcome from this. And also, importantly, can we diagnose a cancer in these patients or do we need to be thinking seriously about cancer? And so, as I mentioned earlier, there's big differences between the cancer frequency and, say, patients with ANNA-1 or Lgi1 and CASPR2.

So having this information up front is important, and so we don't advocate for testing for all antibodies one-by-one for that reason. And then there are-- also we have to remember that there are some more restricted forms of some patients who present with more of an epilepsy or seizure predominant presentation, at least initially.

So thinking about kind of the other potential presentations beyond limbic encephalitis, these are being increasingly recognized, and here's an example of GF autoimmune and GFAP astrocytopathy that was described to Mayo Clinic in 2016. So these patients have a presentation of more like mimics, perhaps tuberculosis or sarcoidosis where you have a lot of leptomeningeal enhancements, but, indeed, these patients have an autoimmune central nervous system disease that's very steroid responsive, and usually not accompanied by cancer but can be in some circumstances. Other examples that are outside of the realm of limbic encephalitis which include NMDA-receptor encephalitis, GABA A-receptor encephalitis where you have kind of more of a multifocal cerebral hemispheric white matter disorder with seizures, and then also the neuronal intermediate filament antibodies.

So an example of how autoantibody testing assistance in our clinical practice is shown here. And so this is an example of autoimmune cerebellar ataxia where a patient presents with rapid-onset dysarthria, incoordination, gait disturbance, on the MRI they may have atrophy of the cerebellum, but on the face of it, we didn't know anything else about this patient. It might be quite difficult, even though we're suspecting autoimmune, to predict exactly what the outcome is likely to be from treatment and also what cancer to look for.

But in the laboratory, we might see that PCA-1 antibody, like I showed you earlier, this anti-Yo antibody that I showed you on the laboratory techniques side, about which I was-- OK, this is likely gynecological breast cancer, but also, unfortunately, the immunotherapy responses probably won't be very robust, and sometimes patients even progress through those treatments to requiring wheelchair. But we might also see something different in the lab, even in the patient who presents like that initially. We might find this one or other of these antibodies like mGluR1 antibody and then Septin-5 antibody.

We described Septin-5 antibody in 2018, and you can see the staining pattern here in the lab, and this is more of a synaptic-type neuropil staining pattern that we see here in the cerebellum particularly. This is distinct in the laboratory to what I showed you earlier that stains largely neuronal cytoplasm. And so the Septin-5 patients generally don't have cancer and also seem to be immunotherapy responsive. And so the outcomes in these patients may be better than what we would see in the traditional paraneoplastic ataxias.

And so here's an example then of how we might build out a profile of autoantibodies that are pertinent to ataxia in the lab. So a patient's serum or spinal fluid is sent from the autoimmune neurology clinic across the street to the neuroimmunology lab, and we could see one or other of these different standing patterns that would lead us to suspect one or other of these particular autoantibodies. And, as you can see here, then we would reflex that then to a cell-based assay in order to ensure that we get the right molecular targets to confirm what we're suspecting. So like what I showed for the limbic encephalitis earlier, we recommend, in this scenario, really taking a broad approach, testing for autoantibodies in patients with suspected autoimmune ataxia.

And here's a list of the various autoantibodies that are important here for ataxia. So for cancer screening then, that would largely be guided by either what's found in the lab by the type of antibody found. So for a small cell lung cancer patient, would probably be a [INAUDIBLE] suspected based on the antibody. And somebody, say, that has had ANNA-1 or anti-Hu detected, would be PET-CT. And then for a patient that has gynecologic cancer suspected in a PCA1 or Yo patient, would probably be ultrasound with transvaginal views or MRI.

So then the treatment approach is largely around trying to establish-- is the person to immunotherapy responsive? What they're likely to respond to, and then what we need to do going forward to try to maintain that improvement or remission. So very often, initially, we're kind of thinking about-- OK, what are the objective measurements that we're going to bring to bear on this situation? And it might be an MRI scan or something in the neurologic exam. And then we're observing to see if there's any improvement in those patients, and then considering an alternative treatment if that particular treatment does not work.

If we do see improvement, then we might think about-- OK, is this sort of one where the person just completely remits and we don't need to do anything further? Or is there a sense that this might be kind of a relapsing course? Do we need to continue with kind of a fairly careful tapering of treatment? Do we need to consider adding in a steroid sparing medication for those patients who need longer term treatment? And then also thinking about some of the more novel drugs such as eculizumab that we mentioned earlier in neuromyelitis optica.

So another thing that's very important in this situation is the time to treatment. So, again, the importance of recognizing these autoimmune disorders, doing comprehensive evaluations quickly to try to establish the diagnosis, and then also initiating treatment quickly. And so we have noted over time that, really, time is brain in this situation. Not in the hyperacute sense of stroke, but, certainly, kind of over the course of weeks, and it can really make quite a big difference, how quickly a patient is treated.

And then we can kind of see some things here. Some examples of objective measures by MRI and showing those improvements and how that can help. So when a patient, say, with GABA A receptor encephalitis, you see improvement in the T2 signal abnormality, in the temporal lobe. And then you have similar things here then for GFAP astrocytopathy, where you have autoimmunity and inflammation in the brain, in A1 and A2 and the T2 flare and the post-gadolinium sequence. And then you see the resolution, but then when the steroids are tapered in this particular scenario, the patient relapsed, and you can see it here radiologically. But then they were treated with high dose steroids again, and managed to have a remission and be maintained on the steroid-sparing treatment only, this mycophenolate.

So the purpose of this site is to demonstrate that there are ways of predicting outcome, and this is an example of a scale devised by Dr. Div Dubey in our group, whereby he devised this APE score based on a variety of different factors that the patient would have a presentation, both clinical and under testing, including their antibody test results, what the likelihood of the patient's seizures improving with immunotherapy. So this can be helpful because when patients have intractable epilepsy and there's a consideration of-- OK, should this person be a candidate for brain surgery to try and treat this? Or for some kind of neuromodulation or stimulation of the brain, often patients are referred to us for consideration of-- OK, could this possibly be something autoimmune? Should we be thinking about immunotherapy? What's the likelihood of an immunotherapy response?

So in summary, these autoimmune CNS disorders, they're important to consider because these do occur in clinical practice. So, collectively, they're more common than first believed, as demonstrated by some of the epidemiologic studies that have been done. They're potentially treatable, but also may be indicative of occult cancer. And the clues may emanate from history exam, serum and CSF testing, evaluations, and response to treatment. And I hope I'm showing you how collaboration between a clinical diagnostic and development laboratory and a physical face-to-face clinical experience can really bring a lot to bear in terms of timely treatment, timely evaluation, and treatment of these patients. Thank you.

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