Update on Antiphospholipid Antibody - In-Depth Overview for Physicians

Grand Rounds


Michael D. Lockshin, MD

Attending Rheumatologist, Hospital for Special Surgery
Professor of Medicine, Weill Cornell Medical College
Director, Barbara Volcker Center for Women and Rheumatic Disease


I want to give you both an overview of the antiphospholipid antibody syndrome and update the audience on things that happened at the recent meeting in Tours, France, which gave some of the most current and up-to-date information on the syndrome. I am also going to give you some speculations about the future.

The basic definition of the syndrome has been around for more than a decade now and has consisted of recurrent thromboses, venous or arterial, recurrent pregnancy losses (I will have more to say about that in just a few minutes), the presence of an antibody, a diagnostic antibody within a family of antibodies, not a single antibody, and some people will add another major component which is the catastrophic vascular occlusion syndrome. The clinical manifestations can be very dramatic and they happen in young people. This is one of the first patients I ever saw here at this hospital, probably 20 years ago, before we had a diagnosis of the antiphospholipid syndrome. This is a young lady, 21 years old at the time, who had multiple cerebral infarcts and a known lupus anticoagulant. I have used the term here now, which we did not have then, of antiphospholipid antibody syndrome or PAPS. This also is a MRI, one of the earliest versions of the MRI, demonstrating diffuse white matter disease and infarctive disease in a 33-year-old man whose presenting manifestations were dementia and occipital blindness. This is a more common manifestation; this is a lady whom I saw just about two weeks ago, a 42-year-old woman who had multiple UBOs (or just diffuse areas of hyperintensity) scattered throughout her brain-you can see them along here or in contrast along here-and forgetful episodes with some hints of dementia but very little else as a manifesting symptom. You can also see abrupt and severe vascular occlusion. This was a spontaneous event in the lady who ended up developing gangrene of three of her extremities before the catastrophic syndrome was stopped by plasmapheresis, and who survived this event despite the severity of what happened.

The way in which I got into studying this syndrome was with the pregnancies. This again, is a relatively old slide, demonstrating in a patient population with lupus and primary antiphospholipid antibody syndrome alone, the risk to pregnancy of having the antibody going from no antibody present (under 16 IgG antiphospholipid antibody units) to high titer on the first blood test drawn during the index pregnancy. In the green bars are women who have not been pregnant before. The yellow bars are women who have had prior losses, and you can see that in this group of patients the history of a prior fetal loss and a high titer antibody gave an over 80% probability of losing the current pregnancy. There was stepwise increment from negative titers but it was also true that the history of a prior loss even in women who had negative antibody titers also predicted loss at about a 2:1 ratio, that is, a prior loss plus the high titer antibody. Predicted losses in a woman who had never been pregnant but who had high titer antibody had about a 40% probability of losing that pregnancy. I guess it does not show well on this slide, but one of the characteristics also is livedo reticularis. This, in a better projection, shows rather striking livedo. It is characteristic of many patients and is distinct from a more hard livedo, as you see, here referred to as livedo racemosa, a point made by Jean-Charles Piette from France, that is different and that occurs in patients with vasculitis. There is a good paper by him in the recent Journal of Autoimmunity that describes the differences between the two types of livedo that occur, the one with antiphospholipid antibody and the one with vasculitis. This is one of the severe manifestations occurring in a young woman with the catastrophic syndrome. Those of you who recognize the histology will recognize this as myocardium. This is a coronary artery that has completely occluded and was the cause of her death. Again, I do not remember her precise age, but she was in her thirties. You will note the characteristic pathology, which is a totally bland thrombus like this, with no signs of inflammation, no vasculitis whatsoever within the coronary vessels. This is another patient, a young man who was well known to many of the physicians here. This is his aortic valve, along here, with this huge excrescence that led to severe aortic insufficiency and eventual valve replacement in this man, and also, mitral valve replacement. This is another manifestation of the syndrome that occurs in patients who have had the disease for some period of time. In the catastrophic syndrome, a relatively rare but dramatic event that can occur is the sudden onset of adrenal failure associated with hemorrhage around the adrenal glands. The hemorrhage occurs after infarction of the adrenal glands and is the cause for sudden hypotension, severe nausea, and back pain in patients with the antiphospholipid antibody syndrome; this slide given to me by Ware Branch shows extensive hemorrhage around both adrenal glands.

Well, after the preceding antiphospholipid antibody meeting in Sapporo, Japan, in the end of 1998, a committee put together criteria for the classification of the disease in clinical studies. Those criteria were set up largely by consensus rather than by formal tests and were published in Arthritis and Rheumatism in mid 1999. The criteria that are now used for the definition of the syndrome include clinical criteria (which include thrombosis, one or more confirmed episodes of venous, arterial, or small vessels disease or and/or pregnancy criteria, which is one or more unexplained fetal deaths after ten weeks of pregnancy, that is, after the appearance of the fetal heartbeat, or one or more preeclampsia or placental insufficiencies occurring before 34 weeks - something felt strongly by the obstetrician- or three or more unexplained consecutive spontaneous abortions less than 10 weeks).

In addition to the clinical criteria there are laboratory criteria. The laboratory criteria are medium or high titer, not low titer, IgG or IgM anticardiolipin antibody, to the extent that it can be demonstrated (and I will come back to this) beta 2 glycoprotein 1-dependent, on two or more occasions at least six weeks apart and/or a lupus anticoagulant on two or more occasions at least six weeks apart. There are reasons for that that I will show you. The lupus anticoagulant has definitions that must include a prolonged phospholipid dependent screening test such as an activated partial thromboplastin time or a dilute Russell viper venom time. The definition of prolonged is not made, does not state whether it is one second or 20 seconds. It must fail to correct with mixing, it should be shortened with excess phospholipids such as the platelet neutralization test or other tests that are used, and, in general, you would like to know that there are not other coagulopathies present, although it is not always practical to test for those. Now there are problems with the definitions because not all of the tests are standardized. What is fairly clear based on efforts by Nigel Harris and others is that the IgG and IgM anticardiolipin antibody can be reproducible, defined at least by good laboratories, repeatedly with similar technologies. IgA anticardiolipin antibody is more or less standardized but has never been subjected to a formal international test but in late 2003 is generally accepted as standardizable). A lupus anticoagulant, although the hematologists have had several attempts- several international meetings to standardize lupus anticoagulant-my judgment of that-hematologists may differ-is that they have failed to provide a single consistent definition. Tests for the antibody to beta 2 glycoprotein I, anti-phosphatidylethanolamine, or anti phosphatidylserine have never been formally standardized across multiple laboratories nor has the lower limit of the lupus anticoagulant been defined. That comes back to the issue, if a test is one second over the control, does that count as a lupus anticoagulant? There have been some studies of reproducibility, and these studies are about five years old now and have not been redone to my knowledge, and I would hope that we would have something better.

This is testing nine suppliers of kits to measure the antiphospholipid antibody. The Louisville standards, or Nigel Harris' standards-and this is a relatively high normal of 10 units GPL and a high titer positive of 80 units GPL-tested against several kits that were available in Europe for this study but are available here as well. What you see is a fair amount of variation from kit to kit. Outlined in blue are some kits that absolutely failed to meet anything that resembles standards. Outlined in blue, those that are 50% or more different from the Louisville standards. Please see here that there are two that just read high titer antibody. Here is one that is called a positive, and one that would be called negative by every other laboratory. Here is another one that fails to reach high positive. This is the reason why we speak in the criteria of having repeated tests preferably by different laboratories over a period of time. The lupus anticoagulant is not much better. This was presented by Doug Triplett in Tours, and this is for those people who believe that the dilute Russell viper venom time is the be-all and end-all of the measurement of the lupus anticoagulant. What I have listed here are patients with known lupus anticoagulant and seven different suppliers of dilute Russell viper venom for the tests. Pretty much everyone who has lupus anticoagulant except for supplier G was identified by dilute Russell viper venom time. However, pretty much everyone, by many of the suppliers, who was on heparin was also defined by the Russell viper venom time. A few of the suppliers did not pick up any. Oral anticoagulant showed up as positive for most of the dilute Russell viper venom times. Anti-factor VIII: a fair number also read as positive. Hemophilia A's were picked up by some of the manufacturers with the dilute Russell viper venom time. Fortunately hemophilia B's were not. These gave sensitivities quite high for the dilute Russell viper venom time but specificities really quite unacceptable. I bring this up particularly because it is common for hematologists, in particular, to say that if the dilute Russell viper venom time is positive there is a lupus anticoagulant present. I am just pointing out that that is not so. It is a very nonspecific test.

Now we had the opportunity to test the Sapporo criteria with a population of patients here. Together with Dr. Sammaritano and Schwartzman we gathered together 243 patients with lupus, lupus like disease, or antiphospholipid antibody to test the Sapporo criteria, and what we found was that the criteria worked quite well. All of these criteria here are highly statistically significant in separating primary antiphospholipid antibody syndrome from lupus patients without antiphospholipid antibody syndrome, that is, fetal loss greater than ten weeks, three or more fetal losses earlier than ten weeks, arterial thrombosis, venous thrombosis, high titer IgG and IgM, and lupus anticoagulant. These are the percentages that we saw in our population as positive. It is important to note that lupus patients with no antiphospholipid antibody can have fetal losses greater than ten weeks and, of course, they can have arterial and venous thromboses also. So just the fact of having lupus and that type of problem does not make a diagnosis for antiphospholipid antibody alone. We also tested criteria that were not in the Sapporo criteria. Low titer IgG and IgM antiphospholipid antibody failed to identify patients with antiphospholipid antibody. However, IgA only did identify it, although only 10% of the people with antiphospholipid antibody had IgA. Biological false positive test for syphilis was quite good. Thrombocytopenia, defined as platelet counts less than 100,000 on two or more occasions, also was reasonably good at separating the groups; only 9% of the lupus patients have that type of thrombocytopenia. Livedo reticularis was present in 21% of the PAPS patients but only 3% of the lupus patients. The people who defined the criteria in Sapporo were wise. One or two losses less than ten weeks did not separate the two groups; preeclampsia did not separate the two groups. Alarcon-Segovia from Mexico has insisted that autoimmune hemolytic anemia is a criterion. We did not find that it separated the two groups either, although we had very few patients with autoimmune hemolytic anemia. All of these, in gold here, are highly statistically significant. So the summary of our look at the criteria show that the criteria are quite good at identifying the patients with this syndrome; and biological false positive test for syphilis, IgA antibody phospholipid antibodies, thrombocytopenia, and livedo are also good at diagnosing such patients.

Now it is important to keep in mind if you are faced with a patient with thrombosis or a patient who is undergoing hematological study that there are quite a wide variety of other anticoagulants than lupus anticoagulant. This is a summary also put together by Doug Triplett. I am not going to run through all of these, but these are different ways in which you can get anticoagulants in the blood that are not lupus anticoagulant. It takes a fairly extensive hematological evaluation to study these. There are neutralizing antibodies and non-neutralizing antibodies, including the factor II inhibitor that Dr. Erkan has written about here.

Hypercoagulable states to think about, if you are faced with a patient with hypercoagulability, are as follows (this is something that is often missed, as I see it reported around the country). It is useful to think of them as arterial states or venous states. If you are faced with a patient with hypercoagulability with a venous thrombosis the list is quite large of what you look for: lupus anticoagulant, anticardiolipin (separable), protein C and protein S deficiency, antithrombin III, the factor V Leiden, the factor II mutations, a prothrombin mutation, prothrombin activator inhibitor mutation-all are associated and these lists are being added to every day on the venous side. Arterial clots have very few causes, lupus anticoagulant, anticardiolipin antibody- and I should point out to you that the patients may have one positive or both positive, these are separable antibodies-and hyper-homocysteinemia. Other than that, if you have a young patient with an arterial occlusion and no other obvious reason for the arterial occlusion you are basically looking at that very short list.

Regarding pregnancy loss, the coagulopathies all cause pregnancy loss. This is a recent summary of several of the coagulopathies, antithrombin III, protein C and protein S deficiencies, factor V Leiden. What is displayed here is large number of pregnancies looked at. These are the relative risks of a miscarriage and the relative risks of a stillbirth, stillbirth again being the greater than ten weeks. Antithrombin III, protein S deficiency or combination of deficiencies all gave very high relative risks. This particular study did not compare antiphospholipid antibody patients. They were excluded from this study, but coagulopathy in general is a threat to pregnancy survival. It is also important to note that, if you find incidentally antiphospholipid antibody, there are a lot of other associated causes, some of which are simple viral infections, and so on. Nigel Harris pointed out other patients who had high frequencies of anticardiolipin antibody, looking at tropical diseases: kala azar had a high proportion of beta 2 glycoprotein I antibody, syphilis of course, and Leptospirosis, and on and on and on. I think it is important to keep in mind that infectious disease is quite capable of producing the antibody without thrombosis.

If you have the antibody and are asymptomatic, what is the risk that you will develop an event? There is only one prospective study that has looked at this. This was The Doctor's Health Study that ran out of the Brigham and Women's Hospital and this demonstrated young men, healthy men with blood tests that gave GPL units in these ranges. These ranges were defined by statistical maneuvers. Relative risks of low titer antibody was 1, starting to get positive it goes up, and it goes up to a relative risk of 7.6 for venous thrombosis pulmonary embolism, which was the only thing seen in this particular population, but an important one because these are healthy physicians, male predominantly, and the disease is predominantly female but it still demonstrates a fairly high relative risk.

In terms of pregnancies, there are a number of studies that have been done. This is one that happened to have a fairly large population of women who had been well studied. The definition of pregnancy loss here was three or more losses in the group of patients studied, well women, not lupus, not previously defined as antiphospholipid antibody. Approximately half had no known cause of the pregnancy loss after intense study. Some had anatomic problems: these are predominately bifid uteruses, septate uteruses, and things of that nature. Some had other genetic causes, very few had infections, and in this particular study, which gives a relatively high number, 21% of women presenting with three or more pregnancy losses, had antiphospholipid antibody. That number, I said is relatively high. Most people get it around somewhere between 10 and 15 percent, clearly a sizable proportion of all women with repeated pregnancy loss, but by no means the totality, of explanations for pregnancy loss. This particular study did not look at other coagulopathies.

Mechanisms for these events: There are a lot of hypotheses and a very active research enterprise going on right now. Activated endothelial cells is one of the major thoughts that is currently going on. Jane Salmon has participated in this as have the hematologists Roy Silverstein and Ronit Simantov across the street, and particularly looking at upregulation of ICAM-1. The same sort of work happens in the trophoblast, in which ICAM-1 is upregulated and the antiphospholipid antibody interferes with that or participates in the upregulation of the adhesion molecule. Other work has looked at trophoblast differentiation in pregnancies and that has to do with the in vitro and in vivo evolution of trophoblast during the course of early pregnancy. Lisa Sammaritano here, has looked at the interaction between placenta anticoagulant protein 1, also known as annexin 5 an antiphospholipid antibody, as has Jacob Rand at Mount Sinai (I will have a little bit more to say about that in a minute.) As a mechanism of reducing what is a normal anticoagulation that occurs in pregnancy, which may prevent the rejection of the fetus, reducing that anticoagulant in the placenta allows the placenta to clot. (And a word that I will come to again in a minute from Jane Salmon has to do with compliment regulation within the placenta.) The pathology of placentas has been studied by Margret Magid, who with our population of patients, looking at lupus patients as opposed to antiphospholipid antibody but identifying those who are antiphospholipid antibody negative and those who are antiphospholipid antibody positive for a variety of placenta criteria. The ones in yellow are the ones that were statistically significant: infarcts in the antiphospholipid antibody positive, ischemic hypoxic change in the ones that had antiphospholipid antibody, and the rest of these terms which are rather technical terms for defining placental pathology did not distinguish the lupus negative phospholipid antibody negative from positive placentas. An example of this is a placental thrombosis in a relatively early placenta from one of our antiphospholipid antibody patients.

There are animal models to study antiphospholipid antibody. This is a slide given to me by Paolo Casali and published in Arthritis and Rheumatism in 1998 showing the mouse pregnancy model. This is a mouse uterus with the pups all lined up in a row like railroad cars and these are the pups dissected out of the uterus demonstrating the size for this particular duration of pregnancy. This mouse has been given IgG-non-antiphospholipid antibody IgG. This mouse was given antiphospholipid IgG and has little nubbins for some of the pregnancies and the pups are much smaller along the way as well. So you can measure the amount of resorbed pups, the number of pups that are viable. This is now a very well established model for measuring antiphospholipid antibody. It can be regularly reproduced and it basically causes mouse abortion. In part this is controlled by complement. This is work by Jane Salmon presented last year at the ACR meeting. Looking at the resorption rate, that is the number of fetuses, and the embryo rate, if you give antiphospholipid antibody to a mouse you get about 40% resorption and small fetuses. If you give IgG alone you do not get that. This is a complement regulatory protein that has been made to persist in the circulation. If you give just that complement regulatory protein and no antiphospholipid antibody you do not get much in the way of resorption, or you get less resorption and bigger fetuses; and if you give an antiphospholipid antibody plus that protein you also do not get the resorption rate and you get larger fetuses, implying that complement is intimately involved in the rejections or the absorption, the killing of the fetuses in antiphospholipid antibody syndrome, a relatively new thought in the description of how fetal loss occurs.

There is another model for thrombosis and this is derived from Silvia Pierangeli and Nigel Harris, where they dissect out a mouse's femoral vein, injure it, and watch a clot form. You can measure the clot in terms of its size and you can measure a clot in terms of its duration. You give normal human serum or saline you do not get much in the way of clot size and it does not last very long. If you give antiphospholipid antibody you get larger clots that last for a much longer period of time. This is the other model that has been used to study the antiphospholipid antibody. Using these models, Silvia Pierangeli, and Nigel Harris, and Jane Salmon have looked at a variety of different mechanisms for antiphospholipid antibody and others have looked at this as a measure of treatment protocols. There are some interesting things that have developed. Fc gamma knockout mice given antiphospholipid antibody do develop clots, indicating that is not an important mechanism, but if you have an ICAM or VCAM or a P selectin knockout plus the antiphospholipid antibody, that animal does not develop clots. The implication is that you must have the upregulation of the endothelial system to develop the clot. If you give intravenous immunoglobulin you lower the antiphospholipid antibody and you reduce the clot size, and, further, if you treat the animal with a variety of things, heparin and aspirin and interleukin-3, and on and on, there are a whole host of things that have been tested, you can prevent the clotting. You can prevent the fetal loss. According to recent work by Girardi and Salmon, an intact complement system is absolutely required for fetal killing to occur. Heparin, which inactivates complement, prevents fetal death; hirudin, an antithrombin anticoagulant that does not inactivate complement, does not prevent fetal death.

Another new idea that has come up very recently is the idea that modified tissue factor is intimately involved in the development of antiphospholipid antibody. There are basically two papers that have described this. One looks at a tissue factor stimulation index, in which the authors are stimulating peripheral blood leukocytes, indicating a very high level of stimulation index in patients with antiphospholipid antibody syndrome and not in patients who have antiphospholipid antibody but who have not had clotting, not in general autoimmune patients, and not in normals. This has been cited largely by Rob Roubey, who has his own work suggesting a similar sort of thing. He looked at antibody to tissue factor pathway inhibitor and he found that it is a totally different antibody from that which is reactive with beta 2 glycoprotein I. He found it in 21% of PAPS patients but he also found it in other connective tissue diseases in patients with thrombosis without connective tissue disease but not in controls.

However, the major work that is going on in this field has to do with phospholipid binding proteins and there are a number of them. I mentioned already the placental anticoagulant protein I or annexin V. There is beta 2 glycoprotein I; prothrombin is a phospholipid binding protein; and there are antibodies to prothrombin that can cause a similar syndrome, and to protein C and protein S and tissue factor pathway inhibitor. Looking first at the placental anticoagulant protein or annexin V - same molecules, just two names for it - there are basically three observations that have been made. Lisa Sammaritano pointed out that it increases the clotting time and it inhibits the antiphospholipid antibody reaction in a clotting essay. La Rosa with Meroni in Italy, looking at placentas which we in part supplied, and which they had on their own demonstrated the presence of this antibody normally in placentas not blocked or present in higher quantity in PAPS placentas, but present in normals as well. Then Jacob Rand has taken this to the physiological level and pointed out that antiphospholipid antibody reduces its expression in the placental villi, in cultures, and he has also looked at endothelial and trophoblast cultures, and it increases clotting in a series of physiological assays that he did with the placenta cultures. There is a good clue that this molecule is reasonably important in pregnancy losses and that antiphospholipid antibody can interfere with it. There are some technical aspects of the assay, which make it not absolutely proven but it is a good hypothesis.

Looking at beta 2 glycoprotein I: Beta 2 glycoprotein I was discovered in 1990 as the cofactor, which is required for antiphospholipid antibody to bind phospholipid. This is what the molecule looks like as crystallized by Flip DeGroot and Ron Derksen in the Netherlands and published in EMBO Journal late last year. This is the molecule seen from two different sides. It has five domains in it. The first four domains are more or less alike. The fifth domain is slightly different. It is part of the complement control protein family. The blue represents positive charges, the red represents negative charges, and the white is more or less neutral. It is a "J" shaped molecule at least in one of its configurations. Looking at the fifth domain, which is the active end of the molecule, up closely a good part of the activity depends on all of these lysines scattered through it. That is also the negative charges of the lysines here which are essential for its activity for its binding to phospholipid. If you mutate the lysines or if you break cysteine double bonds you destroy the activity, you destroy the immunogenicity of the molecule. The model that Derksen and DeGroot have proposed is this: This is a molecule schematically now looked at upside down, first, second, third, fourth, and fifth domain. What they propose - this is a cell membrane - ordinarily the negatively charged phosphatidylserines are on the inner leaflet of the cell membrane. When a cell is activated or when it is undergoing apoptosis the negatively charged phospholipids, that is, primarily, phosphatidylserines, flip to the external surface. What the Dutch propose is that is the point at which the beta 2 glycoprotein I binds to the cell membrane and performs various physiological functions, which I will come to in a minute. So any negatively charged surface will bind to this, but the proposal is that the phosphatidylserines appear only on activated or dying cells.

The origin of the antibody to the antiphospholipid antibody is putatively infection. Looking at the very active component of the fifth domain of beta 2 glycoprotein I: What I have outlined for you here is the amino acid sequences of a variety of peptides, pointing out all of the lysines at this end of the sequence. This is beta 2 glycoprotein I. It is highly conserved and is very similar in cows, horses, sheep, mice, and guinea pigs, and so on. What Aziz Gharavi did was take other peptides that come from infectious sources, adenovirus, CMV, and B. subtilis all having the same forms of multiple lysines at the end of this domain. The blues are hydrophobic areas and the oranges are the lysines. Gharavi immunized animals with each of these peptides, produced antibody to beta 2 glycoprotein I, and has hypothesized that the infectious peptides are capable of inducing the antibody.

What is its function? No one knows what beta 2 glycoprotein I does in the body, but there are lots of thoughts about it. One is that it increases triglyceride clearance, it binds LPa, it reacts with and is demonstrable within an atherosclerotic plaque, it circulates in complexes with apolipoprotein H, it binds oxidized LDL, and it itself is oxidizable. Now there is a purpose to listing all of these things, because I am going to come to some suggestions about where therapy is going to come, and I advise you to keep in mind in particularly the role that this plays potentially in the lipid metabolism of early atherosclerosis. A physiological role that has been proposed for it (other than what it binds to) is that beta 2 glycoprotein I may be involved in the recognition and removal of apoptotic cells. That has been demonstrated and pointed out by Casciola-Rosen. It may also just simply participate in the recognition and response to activated cells, particularly activated cells in the endothelium-that is Pier-Luigi Meroni's thought. Varaala in Finland has been pushing the idea for several years now that it is involved in inactivation of oxidized lipids. It may be a modulator of inflammation in the atherosclerotic plaque. The idea that Yehuda Shoenfeld has suggested is that it may play a role as an anticoagulant itself, which Rob Roubey has also suggested. However, there are deficient mice and knockout mice and there are deficient human patients who have been identified who have no beta 2 glycoprotein I, and those people are completely normal, as are the mice completely normal, so far. Steven Krilis, who spoke here a few months ago, has knockout mice, which at least up to five months of age are perfectly normal. So it is now felt that there is a redundancy of the function of beta 2 glycoprotein I, or that it is not a critical protein. At least its critical function has not been demonstrated yet. (I do not think I am allowed to take questions until the end, before the end of taping.)

Now treatment: That is what everyone wants to know about and that is obviously the most important thing that I will have to say. Treatment for the clotting episodes: Basically we have only one answer at this moment. This answer derives from what I think is still the best paper written on this (at the time this talk was given) by Munther Khamashta from Graham Hughes' group in London, published in the New England Journal about five years ago, a retrospective study, to be sure, but reasonably large and reasonably well distributed. Looking at patients identified as having antiphospholipid antibody syndrome, not just the antibody, who received no treatment, aspirin alone usually in the form of baby aspirin, which in Britain is 50 mg, warfarin to an INR of less than 3, warfarin to an INR of greater than 3, in both cases with aspirin, and six months after discontinuing warfarin. What you see here are patient-years under observation and the events per year in this group. Almost 30% had recurrent events-they all had had at least one event-if they received no treatment, aspirin reduced it to 20%, warfarin to an INR of less than 3 reduced it to 10%, keeping an INR greater than 3 reduced it to about 1%, and discontinuing warfarin gave 130% likelihood of recurrent thrombosis: Very powerful information arguing for an INR greater than 3 in all such patients. This paper has been criticized for two reasons: For one, the paper did not separate venous thrombosis from arterial thrombosis, and the argument has been made that such vigorous anticoagulation is not necessary for venous thrombosis. Secondly, keeping an INR greater than 3 runs a very high risk of hemorrhage and this has been pointed out by Gonzalez-Trujillo from Mexico in the recent ACR meeting, that is well recognized. In this particular study they were running INRs of 4, 6, 8, and so on when they had hemorrhages, though there is an argument to control it at 3, but nonetheless there is a real risk of hemorrhage, so it is not a trivial decision to anticoagulate such patients. [A paper published in 2003, a prospective controlled, double blind trial of two doses of warfarin, concluded that a target INR of 2.5 is as effective as, and less dangerous than, a target INR of 3.5. This paper by Crowther supercedes Khamashta's and now stands as state-of-the-art.]

Regarding treatment for pregnancy. Again, there is one answer available and that is supported by a series of studies from England and from the United States looking at aspirin plus heparin and aspirin alone and demonstrating that fetal survival rates, in relatively small studies, of between 70 and 80 percent are possible. Women entering these studies have had at least two fetal losses previously and that doubles the fetal survival rate of women treated with aspirin alone. It turns out that low-dose non-anticoagulant doses of heparin (5000 units twice daily--Kutteh started at 5000 and upped it a little bit towards later pregnancy) is as effective as therapeutic heparin, raising some questions about anticoagulant doses of heparin, raising some questions about the way in which heparin might work (and offering indirect support to the idea that heparin acts through complement rather than through anticoagulation). There are no formal controlled studies with low-molecular weight heparin, but most people are using low-molecular weight heparin for its convenience and safety right now. For treatment for pregnancy loss aspirin plus heparin throughout the pregnancy is a standard at the moment. Some people are arguing that all the damage occurs in the first trimester, and that anticoagulation could be discontinued after the first trimester. There are no formal papers that state that to present. It has been tried in countries outside of the United States, and we will hear more about that later; but I do not have an answer, and my patients at the moment stay on heparin throughout the pregnancy.

Now something rather startling came up with the study that Doruk Erkan did here and also presented to ACR last month or a few weeks ago, and that was looking at the issue of patients who have not had clots. What is necessary to treat them? This is a retrospective study. The patients who are identified were those who had had pregnancy losses but no thrombosis whatsoever, and Doruk did this by getting patients also from St. Luke's/Roosevelt and from NYU and Beth Israel-Hospital for Joint Diseases together. There were 63 patients, in this particular analysis of the data, in whom there was adequate information available. About half of them, by their physician, had been told to take aspirin on a daily basis and about half had not. Over a ten-year follow-up but actually at about four years about 10% of the women taking aspirin had had a thrombotic event but 63% of the women who were not taking aspirin had a thrombotic event, suggesting in fact that, effectively, asymptomatic antibody carries a very high risk for thrombosis and that aspirin is in fact very protective. There will be a formal study, a prospective study, to verify this but it is startling information at least from my point of view.

There is another bit of information that also came out of this. We had a medical student, who is now a resident in OB-GYN across the street, who looked at women undergoing ovulation induction-in vitro fertilization, who get very high hormone levels, both lupus patients and PAPS patients. The single important relevant conclusion that came from that is that there was no increased flare in these women, for the lupus patients, and none of them developed thromboses, suggesting that hormones themselves do not control the thrombotic events in this group of patients.

Now the future: We are limited at the moment basically to aspirin, heparin, and warfarin. We do not give warfarin during pregnancy in this country because it is consider to be teratogenic, though there are some others who are arguing that warfarin is usable after the first trimester. However, anyone who is treating such patients knows that this is a rather unsatisfactory way; it is highly dangerous, it is very inconvenient, and is not perfectly protective. This was the result of a MEDLINE search I did of what is going on in the vascular world, and I do not intend to walk through all of this, but if you look at interventions for endothelial activations that the cardiologists and for that matter neurologists are looking at for early intervention or prevention of strokes and coronaries, where a huge amount of the activity is, there are huge numbers of things being looked at as potential targets for therapy. This could go on. When I did the MEDLINE search over 500 articles came up. I stopped at about 200 for looking at things that are being looked at. Some of these are here: complement for Jane Salmon, selectins, and integrins and so on. Lots and lots of potential targets, I can tell you that not a single laboratory dealing with antiphospholipid antibody patients in the world is looking at any of this stuff at the present time, and that is an important consideration. You can look at lipid control and antiplatelet agents for prophylaxis. There is something happening in this field at least from Italy where Pier-Luigi Meroni has been interested in lovastatin or Lescol and endothelial cell activation. This is in vitro stuff at the moment but he points out that it inhibits endothelial cell activation whether it is caused by antiphospholipid antibody or whether it is caused by other activators. If you give mevalonate to counteract the lovastatin effect it reverses the drug effect or at least partially and in a mouse model mechanism, unstated at the present time, it increases the MRL-lpr mouse's survival from 159 to 190 days, just presented recently, suggesting that the approach of looking at antiphospholipid antibody as an atherogenic or as an endothelial activation problem as opposed to a coagulation problem may be a valid approach in the future.

The other things that are being looked at, at the moment: Virus products. This is based largely on Gharavi's work looking at those peptides to see if they can be used as immunogens, looking at the beta 2 glycoprotein I peptides as well and looking at histocompatibility complex antagonists on the idea that this, like most autoantibodies, is highly dependent on HLA. I put down novel therapies though they have been used already for some time, intravenous immunoglobulin is used, it is used in pregnancies, it is used in the catastrophic syndrome, and a variety of other reasons. There are a number of potential explanations of how it works. Anti-idiotype therapy is something that comes out of Yehuda Shoenfeld's work. Absorption columns are being attempted by La Jolla Pharmaceuticals to remove the antibody as well.

What do we need to do for the future? Well we need considerable more about definition of what the disease is. Is it ongoing or is it episodic? The assumption is that we have to deal with episodic thrombosis, but a lot of people, myself included, are arguing that in fact it is always ongoing that is why you have livedo, that is why you have low-grade thrombocytopenia, and that the episodes of thrombosis are just sort of when it reaches a specific peak, and perhaps the patients should be considered as having constantly activated endothelium, then we need to deal with that. What should we do about asymptomatic carriers of antibody? Well an argument is being generated that they should be treated, though how they should be treated is unclear, and I hope that Dr. Erkan's prospective study will gives us some answer to that. Long-term outcomes: I have not spoken much about this but in fact we are desperately concerned about both mental function and cognitive function in long-term carriers of this and certainly about the cardiac valvular disease we are running into over and over and over again, having had four patients whose valves we replaced in the past year here for aortic and mitral valve insufficiency, a long-term sequelae, and other interventions as I have been mentioning, largely interventions that we learn about from the vascular disease community rather than our own and the beginning of treatment trials. We are part now of a consortium of antiphospholipid antibody that was organized by Rob Roubey, which will allow us the opportunity to test things on a national basis as well as a local one.

I thank you very much. I think I can take questions now.

Question/Answer Session Following Grand Rounds:

Question: Considering the relative frequency of Factor V Leiden and some of the other mutations, has anyone ever looked to see what happens if both APS and one of those occur in the same patient? And might that explain the catastrophic syndrome?

Answer: The answer is that it does not explain the catastrophic syndrome. There are a number of people who have looked at the simultaneous occurrence of not only factor V Leiden, but multiple events, and the results are kind of mixed. Regarding factor V Leiden there are papers both saying yes it increases the risks of thrombosis and no it does not. In the pregnancy world the data to date suggest that having two hits makes you at higher risk than having one, having three hits in an occasional patient makes it worse than having two. But on the thrombotic risk I think the jury is still out.

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Question: What is the relationship between beta 2 microglobulin/amyloidosis and antiphospholipid antibody syndrome? Do APS patients develop amyloidosis?

Answer: There is a confusion there and this is one I run into in the laboratory all the time. It is not beta 2 microglobulin, which has nothing to do with this disease at all, but beta 2 glycoprotein I. For anyone who is watching this on the WEB who tries to order an antibody to beta 2 glycoprotein I, you will find that the laboratories about 80% of the time will send you back a beta 2 microglobulin. You have to be very specific and make sure the laboratory knows what you are doing. There is no information on beta 2 microglobulin or amyloid or anything else with this particular syndrome. It is a different protein with a similar name.

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Question: Is there any correlation of the antibody titer with the risk for thrombosis and do you follow the titers? With patients who give a decrease do you consider stopping therapy?

Answer: It is an important question and the quick answer is no with an explanation. There is a statistical relationship with titer of antibody. Generally those people who have problem with disease, that is, either thrombosis or pregnancy loss, are high titer antibody patients. Most of the time the lower titer patients are not the ones who get into trouble but there is no direct correlation with titer and it is not useful to follow the titer. There is a publication by Ron Asherson a few years ago and I think Ron Derksen also stated this: That even in those patients in whom the antibody goes away, once they have started thrombosing they will continue to thrombose, so the argument at the moment is that they should remain on continuous anticoagulant therapy. Sometimes it is not practical but that is the official line right now.

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Question: Mike, the patients who have an infection and develop antiphospholipid antibodies, how long will they last after the infection is gone or do they stay there forever?

Answer: That is an important point. The question if you did not hear it was - The infectious antibody how long does it last? That is one of the reasons for doing the two tests six weeks apart. Usually things like CMV or mononucleosis are something that will induce the antibody. It will be a transient antibody that will have disappeared within a few weeks or months. I cannot speak for patients with kala azar, which is clearly a long-term infection, and in Brazil lepers have long-standing antiphospholipid antibody. What was important about that slide, which I did not emphasize, was that an earlier version of the information was that you could distinguish infectious antibody from noninfectious by demonstrating its dependence on beta 2 glycoprotein I, which can be done by a few laboratory tricks or manipulations.


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