Intravenous Immune Globulins in the 21st Century: Progress and Challenges in Efficacy, Safety and Paths to Licensure Lister Hill Auditorium National Institutes of Health Campus Bethesda, Maryland April 13, 2005 PARTICIPANTS SPEAKERS: Paul Aebersold, Ph.D. Don Baker, Ph.D., Mark Ballow, M.D. Melvin Berger, M.D., Ph.D. Rebecca Buckley, M.D. Erwin Gelfand Basil Golding, M.D. Jonathan C. Goldsmith, M.D. Mary Ann Lamb Hans D. Ochs, M.D. Judi Miller Joshua Penrod, JD, LLM, MPH Joan Robertson Richard Schiff, M.D. Dorothy Scott, M.D. Mark Soucie, Ph.D. E. Richard Stiehm Robert P. Wise, MD, MPH CONTENTS Opening Remarks - Dorothy Scott, M.D. Perspective on History of IGIV, Current Important Problems and Questions in the Field - Hans D. Ochs, M.D. I. IGIV Efficacy: Infections in PID Patients: Prevention of End-Organ Damage; Need for Practice Guidelines and Screening - Rebecca H. Buckley, M.D. Surrogate Markers for IGIV Licensure - E. Richard Stiehm IGIV for Primary Immune Deficiency: Antibodies Against Potentially Problematic Pathogens - Dorothy Scott, M.D. Are we Adequately Replacing Antibody in Patients with Primary Immunodeficiency? - Richard Schiff, M.D., Ph.D. Panel Discussion Safety and IGIV Product Differences, - Erwin W. Gelfand, M.D. Safety of IGIV Infusion-Related Adverse Events, - Mark Ballow, M.D. An Emerging Issue with IGIV Products, - Donald Baker, Ph.D. Comments - Marcia Boyle FDA Safety Surveillance for Licensed Biological Products, Including Intravenous Immune Globulins, - Robert P. Wise, M.D., MPH Utilizing Public Health Surveillance to Monitor Adverse Outcomes of Blood Product Therapy, - Michael Soucie, Ph.D. Perspective Post-Marketing Surveillance of Octagam, - Judi Miller Workshop on Intravenous Immune Globulins in the 21st Century--Product Tampering: A Case Study, - Joan Robertson Panel Discussion III. IGIV Licensure for Treatment of Primary Immune Deficiency: Immune Globulin Intravenous (Human) - Basil Golding, M.D. Industry Perspective: Current Clinical Paradigm for PID Indication - Mary Ann Lamb Regulatory Requirements for Subcutaneous Ig for PID - Paul Aebersold, Ph.D. Results of the First Clinical Trial - M. Berger, M.D. Critical Path Initiative - Dorothy Scott, M.D. The Critical Path: The Plasma Industry Viewpoint, - Joshua Penrod, JD., LLM, MPH PROCEEDINGS Opening Remarks DR. SCOTT: Good morning. I think we will get started. First I would like to get through some housekeeping issues but I want to thank everybody for your attendance. I think it will be a very exciting workshop.These are the announcements that must be made: Most importantly, no food or beverage is allowed in the auditorium. I think there is an enforcement arm here so you need to be careful. They want us to remove all food and beverages. Set your pagers and cell phones to vibrate. To activate your microphone, press the mike button. There is a message desk. That phone number is 301-496-4062. People can call in and leave you a message there. We are going to be asking our speakers for permission to post their slides on the FDA web site, and we will also be posting a transcript of this meeting. Now I will get to the more interesting part of this. I want to thank everybody for coming and I want to point out that we have some leading authorities, in fact many of them as speakers and panel members in the PID and IGIV field. We would also like to welcome the many manufacturers and sponsors that have come to provide their input. We have here today people from FDA, including our Office Director, Jay Epstein. We have CDC, CMS and many other people that, we are very grateful, have been able to come. I would also like to thank the Immune Deficiency Foundation for co-sponsoring this event and for providing us with the connections that we needed to help get this agenda and these wonderful speakers. Ms. Marcia Boyle, who is the Chairperson and CEO of the Immune Deficiency Foundation, is here and she may say a few words after lunch. Now, the outcomes and goals of this workshop are to identify the most important current issues in IGIV treatment for people with primary immune deficiency. What we would like to do, among other things, is identify a possible research focus that would be important both in the laboratory and clinically; and to ponder potential solutions to current issues and problems that we have identified. I would ask everybody, please, to participate in the discussions that we have. I think it would be very useful and it will help us to generate some outcomes. Now I would like to introduce our keynote speaker, Dr. Hans Ochs, who is a Professor of Pediatrics at the University of Washington, in Seattle. He has devoted his career to the study of primary immune deficiency and to the clinical care of the people with primary immune deficiency. He has been present at workshops and speaking since time immemorial. I couldn't go back far enough through the workshops to find one where he hadn't participated. We are very pleased to welcome him, and thank you for setting the scene for the entire workshop. Perspective on History of IGIV, Current Important Problems and Questions in the Field DR. OCHS: Thank you, Dr. Scott. That makes me feel a little overdue for the next phase of IGIV research; this morning in the taxi I heard that IGIV was successfully used to treat Alzheimer's, so I am in line for preventive therapy. I have the task to provide you with some retrospective ideas about IGIV, some of the things we are doing now and some ideas that we will have to work on today to create a better future for IGIV. This slide is just intended to point out that the idea about passive immunization is spanning two centuries, going back to the 1890s when Behring and Kitasato found that serum has something that prevents the effect of diphtheria and tetanus toxin. As we go down the line, we learned that the antibody activity is in the gamma globulin fraction. For that, we had to have a way to separate proteins in the serum. We learned how to fractionate plasma, creating Cohn Fraction II, which is the basis of IGIV. Bruton discovered that there was a disease where gamma globulin was required for treatment, namely, X-linked agammaglobulinemia. Then, one of my favorites, Silvio Barandun, in the '60s, started to give gamma globulin intravenously with very interesting results--clinical studies that would not be possible today. Then, in the '80s we learned how to manipulate Cohn Fraction II properly to make this gamma globulin useful for IGIV injection. This is one of the hallmarks of the creating of IgG product, finding that the antibody activity is in the gamma globulin fraction. Very simply, plasma was separated based on the charge of the molecules and the size of the molecules. Around the same time, the secret of IgG was revealed, knowing now that these molecules have a heavy chain and a light chain; that there are variable regions; that there are constant regions which are the different genetic control and are the different isotypes. We also learned on the way how nature is putting together this incredible variability of these antibodies by putting together Vt pieces and then, on top, that there is a possibility to change these things somatically due to somatic type of mutation which is not genetically determined. So, we have a final output that can adapt to almost any foreign protein or polysaccharide. That is Silvio Barandun at the time when he used Cohn Fraction II for intravenous use, and it is striking to read these papers of his. He would take Cohn Fraction II and inject it in normal controls, and he found out that 1/10 would have an anaphylactic reaction. If he took patients which looked retrospectively like common variable immune deficiencies, he would give this Cohn Fraction II in very small doses and 9/10 would have an anaphylactic reaction. He also learned--and this was very important for me later on--he found out that patients who underwent this anaphylactic shock were refractory for 3, 4, 5 days. So, when we did the first intravenous studies we actually primed the patients with a small dose of these not very well designed intravenous preparations, and then the next day they could tolerate the material very well. This also indicates to me that more frequent infusions of IGIV are safer--let's say once every 2 weeks instead of once every 4 weeks. So, at the beginning of our journey of IGIV some principles were put down which are now common ideas: highly purified immunoglobulins with no preservative; of course, no infectious agents and we learned the hard way that we actually infected some of these patients with hepatitis C. It should be monomeric or dimeric, not having many aggregates, and biologically active. IgG subclasses should be distributed relatively normally and then, of course, we need to have a broad spectrum of antibody activity, which we have essentially created by using many units for one batch of gamma globulin. This is the result of one of the first IGIV trials in the United States. It was a product from Hyland and we called it modified immunoglobulin in glycine. They called it modified IgG. Then, after seeing this huge amount of side effects, with 76 percent of all patients having relatively serious side effects of pain and fever and chills, and more than half of the infusions were associated with side effects, a simple step was taken, adding maltose to keep the aggregation from occurring and that reduced the side effects dramatically. I have another slide showing this, during the same study. If you take the number of infusions with side effects--this is the modified immunoglobulin in glycine and the red one is in maltose, 10 percent maltose--and all these reactions markedly disappeared if the sugar was added to this preparation. This was in the lab. We take Cohn Fraction II at 20 degrees and just keep it for an hour or half an hour at room temperature. Then, if you take this Cohn Fraction II, which was the intramuscular gamma globulin, and heat it for half an hour at 61 degrees Centigrade, which is not usually aggregating protein but in this situation you got a significant aggregation of IgG. If one added to Cohn Fraction II maltose at 10 percent, you could totally prevent the aggregation of protein, and the same thing happened if one used the mitigated IGIV with glycine and added 10 percent maltose. It also withstood this trauma at 61 degrees. When we looked a little further, this is the optical density--you know, the "Milky Way"--and this one is the time at 61 degrees. This is all Cohn Fraction II. If one takes Cohn Fraction II as it is and incubates it, it takes a long time, almost half a day, to get significant aggregation. If one adds methanol, a small amount of polyethylene glycol, this increases markedly this aggregation at optical density of 660 nm. If one takes the same preparation, Cohn Fraction PEG or methyl alcohol, and adds 10 percent maltose this aggregation is almost completely inhibited. Now, what is the therapeutic action? Today I think we are mainly talking about replacement therapy but IGIV also has anti-inflammatory action. It can induce Fc receptor blockade which we use for IGIV in autoimmune diseases. There is an anti-idiotypic autoimmune phenomenon. There are anti-idiotypic antibodies in IGIV. Then, of course, it has an effect on complement components. The primary use for my field of interest and of today's meeting is IGIV in primary immune deficiency diseases, mainly those which have a predominant antibody deficiency like X-linked agammaglobulinemia as the hyper IgM syndromes which consist now of at least five genes. One of them is a T cell-related gene. This is CD40 ligand which also has an effect on the production of immunoglobulin common variable and then IgG subclass deficiencies if it is associated with antibody deficiency. Then, SCID patients either before bone marrow transplantation or some after bone marrow transplantation, if a B cell defect is remaining will depend on IGIV. This was one of the early attempts trying to figure out how to treat these patients. If one gives the old dose of 100 mg/kg, as is shown here, once a month that is not enough to raise the peak nor the trough level. If one gives 200 mg/kg every month one gets a modest rise in the peak level and also a rise above the baseline of the trough line. If one gives 400 mg/kg once a month one gets a further rise in the peak and a further rise, and it levels off later, of the trough level. But you can see this huge difference between peak and trough level. If one gives 200 mg/kg every 2 weeks twice a month you get a much more stable level with less of a difference between peak and trough. If I would give it every week, like 100 mg/kg for a total of 400, you would get even a more steady line. What actually happens during the interval between the infusion and the trough level is a complex way of intrinsic IgG, which may not be worth much, or material from old infusions, the last infusions, and this infusion and so that will determine what actually the peak level and the trough level in a specific patient will be. The serum IgG concentration depends on a number of factors. One, of course, is how much milligrams/kilogram per week, 2 weeks or 4 weeks you give. The route of injection--intravenously you get these high peaks, low troughs; intramuscularly it is painful but it guaranties a more level IgG serum concentration; subcutaneous, which has recently been propagated, is a way to have a very stable serum IgG level. The frequency of infusions, once a month or once a day--the peak level is important because you determine the trough level. The catabolic rate may be different from patient to patient or within a patient. If a patient has an acute infection or is malnourished the catabolic rate will go up. Losses, specifically in the bowel due to chronic bowel disease; and then hydration and also the half-life of a given preparation which we now always determine by pharmacokinectic experiments. I wanted to show you the impact of immunoglobulin on this family which is from Montana. This is a lady born in 1892. She is probably a carrier because one of her daughters is a known carrier on whom we actually did some molecular analyses. Then, two of her sons died in early infancy of infections. This lady has had three sons. One was born in 1944. He died at the age of 4 and did not have the luxury of getting treated properly with penicillin. He died of pneumonia. The next two sons were born in '47 and '49, way after penicillin was available. Both survived but they were not treated with immunoglobulin intramuscularly until they were teens and so they have both had lung surgery. They both had severe bronchiectasis. This patient, I will show you later, developed ECHO infection, ECHO encephalitis, and was the first one treated with high dose IGIV. This one is doing well now. The only complications he had at the age of 43 was cancer of the colon, which is not unusual in patients with X-linked agammaglobulinemia. This gentleman had a sister. She had a young son in 1968. He was born at the time when I saw his two uncles. He was diagnosed very early and is doing well. This is a carrier female, of course, of this gentleman. She has a son. He was diagnosed at birth and both of those are doing very well on IGIV. Actually, this one was placed on intramuscular and subsequently on subcutaneous. He is now in his 30s. That is this individual. I wanted to show a little bit more about this patient, who developed, at the age of 25, an ECHO infection with slowly progressive CNS symptoms. He had all these clinical findings with loss of cognitive skills. He got paresthesia. He had seizures. He had a peculiar dermatomyositis-like phenomenon of indurated skin and muscles, and this was due to a myositis/fasciitis and these patients have an increase in CPK, alkaline phosphatase and transaminases due to loss of muscle mass. The impact of IGIV was quite striking. In 1990 a survey was done of 34 centers and this identified 248 XLA patients. The incidence of ECHO infections before 1985, when IGIV was being introduced, was 39. It is a rough estimated. After 1985 there were only 4 infected. Three were atypical and only one had not been on IGIV by the time he developed this disease. I think it was a patient which Dr. Stiehm knows about. After he developed this ECHO infection this diagnosis of XLA was suspected and he was subsequently treated. There you see this infiltrate of lymphocytes, T cells into the fascia, and the viruses are distributed in the muscle and the fascia. This first patient was treated. He developed this disease just about a year after the first IGIV preparation was tested in the laboratory, and he was treated with more than one liter of IGIV and his symptoms completely disappeared. Safety--of course, we do not want to have any infectious agents in the material. The only one I am really aware has caused problems is hepatitis C. HIV we, fortunately, avoided. Parvovirus is probably neutralized because there is a lot of IgG antibody, and the question of prions is still not completely resolved. We do not want aggregates. The pH should be a little on the low side. Osmolality is really an issue in some patients who have either chronic renal disease or who have cardiovascular problems. Low IgA--this is a question. I don't think many patients have actually a problem with this. Limit the rate of infusion, that means you should not, I think, overdo the amount of IgG per hour. Then, if there are problems we just reduce the interval between infusions and that usually takes care of these general problems. The safety is pretty well now under control with antiviral steps, at least three per preparation. There is viral partitioning during the cold ethanol fractionation which has made this material safe from HIV. There are antibodies are in the IGIV that would, hopefully, neutralize remaining viruses. Low pH is helpful. Beta-propiolactone or caprylate treatment has been used. Solvent/detergent; pasteurization and, most recently, nanofiltration and UV light has been used to reduce that problem with antiviral steps. Adverse events are still a problem. There are general reactions, especially during the first and second infusion of new patients. There may be anaphylaxis, which is extremely rare but it can happen, especially now if you have the subcutaneous Cohn Fraction II if it would inadvertently be given intravenously. Acute renal failure is a problem probably due to high osmolality. Then, there is a problem with the cardiovascular system, volume overload and that can lead to thrombotic events. We have seen a few cases of Coombs positive hemolytic anemia. In fact, one or two preparations, when tested in clinical trials, had positive Coombs test for 1, 2 or 3 days without hemolysis. Then, of course, the CNS aseptic meningitis which is probably also due to overload in terms of osmolality. I just want to point out to you that these general reactions are almost always during the first infusion of CVID patients or when patients are infused when infected. These are to some extent rate related and infusion interval related. As I mentioned, if the interval is too large these patients accumulate--this is a hypothesis--accumulate immune complexes and will have problems. I don't want to go into the details, but for the first infusion we do very careful monitoring. We monitor the rate of infusion of the material. The dose is usually 400 mg/kg and we measure carefully vital signs. We pretreat all these patients when they get their first infusion with Tylenol. Then we have infusion rate that slowly goes up and if there are reactions we have two ways to treat this, one, with SoluCortef and the other is Benadryl. We insist that the first infusion is done in a center where we know how to take care of these infusions. The second infusion is pretty much the same and then they are on their own. So, for the subsequent infusions we do we always use, or most of the time we use 400 mg/kg every 4 weeks or 300 mg/kg every 3 weeks, and our favorite dose is 200 mg/kg every 2 weeks, especially for those who do self-infusion at home. It is no problem and that is the way we are recommending to treat. How do you dose? You have to know the baseline and you want to have a trough level that is at least 300-500 mg above the baseline. Then you may have to increase the dose or shorten the interval if any one of those events occurs, if the IgG level drops or if there is progressive pulmonary disease, or if the patient complains about having the "pre-infusion blues" and they lose interest, they are tired and sometimes they have infections. If they gain weight we have to adjust the dose, or if there are any problems with other adverse events. Now, which preparation should we use? Is there any difference between preparations? There are low dose and low concentrations like 3 percent, 6 percent. There are high concentrations. People are talking about 16 percent intravenously but certainly subcutaneously or intramuscularly that is a good preparation. Should it be lyophilized or liquid? What is the osmotic load due to the formulation? We should look at viral inactivation steps and antibody titers. Source versus recovered plasma is an issue and how many units of plasma are used for a single batch of IGIV. Again, as I mentioned before, the dose you use every 4 weeks or 75 percent every 3 weeks, etc. Should one infuse in a center or at home by a nurse or at home by the patient or a representative of the patient's family. Then, we have to factor in the cost of this treatment. Is there any difference? And, what is the difference in quality of life. There are now interesting studies that address these issues. Self-infusion at home is good for selected patients. It seems to me that is more common in the west than in the east of the United States. The positive aspects of that are that the patient becomes a partner. He can do potentially flexible dosing if it is a university professor; frequent infusions, every 2 weeks, there are essentially no adverse events. The overall events are 0.7 to 0.8 percent, studied by Brennan et al. There is clearly a quality of life improvement. The negative is that there may be lack of supervision that has to be worked out. There may be missing of compliance and one may also miss complications, and in some areas there are potentially problems with legal complications. That was our first attempt to teach these patients. These are all patients in 1981 in the clinical research center of the University of Washington. This gentleman was self-infusing himself. He taught these patients very effectively. The youngest patient who managed to do self-infusion was 6 years of age and he wanted to be as good as his 10 year-old brother, and this family is still self-infusing. There are some rules. Never do it all by yourself, etc. It all makes sense. They have to be very carefully screened every 6-12 months at a center where they are known and where these tests can be carried out so that we do not lose them somewhere in the boonies. The next self-infusion that is on the line, and that will be done in a second, is subcutaneous infusion. In Sweden, for instance, over 90 percent of the patients do self-infusion at home of subcutaneous immunoglobulin. In Britain about half of the population; in the United States the clinical trials have been completed. I have about 10 patients on subcutaneous infusion. That is, of course, very easy. You don't have to find a vein. There are essentially no adverse events, 106 per 27,000 infusions in Sweden. In Sweden it is less costly. The problem, of course, again is supervision and the local reactions that seem to disappear over time with use of subcutaneous infusion. That is one of the early clinical trials. This is on a dose of 200 mg/kg every 4 weeks IV. Then you increase to 400 mg every 4 weeks. That brings you up to a good peak and trough level. In this particular study one last dose was given of intravenous immunoglobulin to measure the half-life and the area under the curve. Then a last dose of IGIV was given followed by weekly injections of 100 mg/kg, practically the same dose as here. Then, after a washout phase the area under the curve was determined. In order to equalize these one has to actually give more, about 130 percent of this. In Sweden they use this dose and they come up with very good clinical results. So, probably one does not have to change the dose. So, to do this effectively one has to have in place instruction on how to do self-infusion. One uses 16 percent Cohn Fraction II as a dose per week. The abdomen or the thigh or the upper arm can be the site of infusion with a small butterfly needle. One can give locally EMLA cream. One can do it with a pump. The best is a syringe driving pump. The infusions take approximately one hour. There is a range. One can use more than one site. There is no premedication and it is okay for infants. As a matter of fact, you can do this by direct push without a pump once or twice a week. I have adult patients who inject themselves every day with the appropriate dose, without any problems, over 2-3 minutes. How do we monitor? In practicality we always ask the patients to write down the lot number. They have to be seen every 6-12 months for follow-up that includes now even a CT scan yearly. That is true for all of our patients. I don't want to go into details but there are a number of different preparations, either lyophilized or liquid, that have different concentrations of IgG; that have different osmolality. Some of them have sugar or no sugar in it, and they have variable pH. There are two others recently on the market, Flebogamma and Carimune, again liquid or lyophilized, and in the future we will have to carefully look at these different parameters to generate the best formulation for this material. That is Bob Good, who is also one of my heroes, if you wish, and I asked myself what would Bob Good tell us what to look for, and I think he probably would say we have to be absolutely sure that our preparations do not have prions. He probably would also advise us that it is very important for the availability that industry, that FDA, and that patients and doctors form a union and address these issues as best as possible. He would probably say it is not so important if it is liquid or lyophilized as long as it is safe. He would also tell us that osmolality is important, especially for patients who can be hurt by hyper-osmolality like patients with chronic renal disease. He would ask the question how much IgG should we give in terms of concentration intravenously; subcutaneously, it is very clear as high a concentration as possible. What about the sugar? On one hand, it is a very good way to stabilize IgG. On the other hand, it provides hypo-osmolal material. He would ask what is the role of spiking the material? Should we use hyperimmune serum, for instance, against TMV to add this to our immune preparation? On the other hand, should we use hyperimmune serum and take it out of the common preparations? Should we use monoclonal antibodies to spike in the future the common IGIV preparations? What is the choice of infusion? I think he would probably say we have to just let the patients and the doctors decide do we give it IV, subcutaneously or intramuscularly, and we have a choice now. So, for the patients it is getting easier. He also probably would tell us that we should simplify the procedure. Self-infusion is great if the patient can do it at home if that is a possibility. Should we do it every 4 weeks, which is the most unnatural way to do it? Should we give it once a week? Or, should we give it every day, the way we do it essentially? And, how can we save the enormous amount of dollars that are going into this form of treatment? I will stop with this and apologize for going over time. Thank you very much. Any questions? Yes? PARTICIPANT: Dr. Ochs, you mentioned a few things that I would like to ask you maybe to comment a little bit more about. One was the situation where some of the PID patients were getting ECHO encephalitis, and you showed there was a drop after treatment but there are still cases--and I know from the time when I was looking after patients at the clinical center that even with treatment you still get PID patients with ECHO encephalitis. My question is should we be looking more carefully at the titers of the antibody, and do we know what titers are protective? And, once the encephalitis has established, is it difficult to really treat it? Is it more prophylaxis than treatment that is going to be effective? DR. OCHS: There have been cases that occurred during IGIV therapy and I have observed two of them with absolutely good records. It is probably very important about what type of echovirus it is and what the titer of antibody in that particular preparation is. The problem is that there are many echoviruses. It could also be another virus. Polio, for instance, was a problem; Coxsackie virus; there is a JC virus. It is almost impossible to get the companies to check for all these viruses. So, we are in a blind alley there. Originally the virology laboratories were willing if we had the virus to actually titrate out different gamma globulin preparations. With the first person we did about 6 or 7 lots and there was a huge difference in specific neutralizing antibody, and we used the highest. That is no longer available. Even the CDC does not provide such service. So, when I noticed these two cases I just told them to use alternatively two preparations, hoping that one of them would have a high titer, whatever came along. But I have sort of forgotten this, but this is one way to do it if you are worried. You know, it is like in the stock market. You don't want to put your eggs in one stock so you put it in different preparations and just routinely switch. But that is very difficult to do and nobody is actually practicing this. But you are right, it still can happen. There is another disturbing thing. One of Dr. Stiehm's former fellows put together a group of patients worldwide of 14 individuals who developed progressive CNS disease without any etiology, and to the best of our knowledge did not have this enteroviral problem. Most of them died. I think there is one still surviving. We do not know what the etiology is. In some patients it could be this virus that has been associated in some patients, this JC virus. These patients have not been carefully studied. Those who were studied did not have prion disease. So, in addition to the known viruses, there are other events that can happen and we do not know if they could be prevented if we would have the right mixture of immunoglobulins in the gamma globulin preparations. But the rate is much lower and the two individuals I have seen with the development of encephalitis during IGIV therapy, one survived and the other one died. It was interesting that both of these patients years before had ECHO encephalitis and they were consistently treated and they responded to the first course, and years later they presented with the same virus. So, it is probably in their system. I would assume that the lot they received prior to their second infection was low in specific antibody. PARTICIPANT: Can I follow up with one real quick question? You stressed in your talk the fact that if you give the IGIV more frequently, for example every 2 weeks, you are going to end up with higher trough levels. I wonder if you have any data? We have tried to find data which would show that and there are trends in some studies that we have looked at that show that towards the end of the 4-week period there is a slightly higher incidence of upper respiratory tract infection. But do you know whether it has ever been documented that more frequent administration would give you not only higher trough levels but reduce infection rates? DR. OCHS: So, the question was if one gives more frequent infusions, let's say every 2 weeks over every 4 weeks, does one reduce the side effects from the infusions that are often seen following infusion every 4 weeks? Yes, formally I don't think it has been studied but the patients, and I have probably about 20 who do self-infusion at home every 2 weeks, they report no side effects. You know, sort of anecdotally, if these patients are put in trials every 2 or 4 weeks, they tell me that they don't feel as well at 4 and then after the infusion they lay around for a day and have some headaches. But there are no formal clinical trials that specifically address the rate of adverse events if you give IGIV every 4 weeks or every 2 weeks. One more question and then we should stop this. PARTICIPANT: Just a quick question, you showed in your early formulation studies that the presence of maltose minimized the adverse events. Then I notice that some of the new formulations don't have any sugars in them. Also, you showed in those early examples that if they heated there was a tendency to more rapidly form what looks like particles. Do you know if the adverse events were associated with aggregates? I mean, did they look by size exclusion chromatography to see if there were dimers, primers and so forth? Could you comment on kind of those concepts? DR. OCHS: At that time we were less sophisticated and these tests were not done, but I noticed in this first preparation which was handed to me in 1973 by Mr. Hardy, from Hyland, that if you looked at the light it was cloudy, there were so many aggregates. So, we used filters and that didn't improve it. Then I noticed that when you dissolved it, rather than in normal saline, in 5 percent glucose the patients tolerated it better. That was the extent of our sophistication at that time and I wish we would have done these other experiments. Now, you can get the same effect by lowering the pH. I didn't show you this but if you go through the motion of adding polyethylene glycol or alcohol and heated you can prevent this if you keep the product at low pH. That was a trick used to have a low pH. They also had another trick, they put a little pepsin in it. So, they were regular alchemists at that time, the Swiss Red Cross Prof. Hessick and Barandun, and they mixed it together right away and when they presented it to the clinicians it worked. But I think the newer formulations, for instance the new preparation that ZIB-Behring has, it has amino acids as a buffer. It took them a long time. I have known about this product for years while it was in development. But they have it pat and they did all these studies which you suggested, and there are no aggregates; they have either monomers or dimers. So, one can eliminate the need for sugar if one uses a good chemist. DR. SCOTT: I just need to announce that the enforcement arm has asked me to ask people really not to eat because the staff here will get in trouble if the management catches you eating, and please take your trash out. On another note, I think there are a lot of interesting issues to discuss and we will save some of those for the discussion section but I appreciate, Dr. Ochs, that you brought up the question of how to optimize dosing frequency and dosing amount because I think that is one of the still current issues. For our session we have Drs. Buckley, Stiehm and Schiff to speak. I don't have time to introduce them thoroughly but if you look at your handouts you will see that we have a brief biography. Just to say that these are all very eminent people in the field and we appreciate that they have been able to come and speak to us to bring up these issues. After the break we will have the panel session and I would also like to invite Dr. Ochs to come up for that. I. IGIV Efficacy Infections in PID Patients: Prevention of End-Organ Damage; Need for Practice Guidelines and Screening DR. BUCKLEY: Thank you very much, Dr. Scott, and thank you for inviting me. I was asked to speak about infections in patients with primary immune deficiency diseases. I will just say at the beginning that IGIV isn't necessarily the treatment that works for all these types of infections because there are certain host abnormalities in certain patients that define their susceptibility to infection and you can't cure that with IGIV. However, the organisms that are sort of characteristic of these other types of infections would be things that people who design IGIV products in the future might want to keep in mind, particularly if you follow Dr. Ochs' suggestion of spiking IGIV with monoclonal antibodies to certain infectious agents. Just to give you an overview about primary immune deficiency diseases, this field, since Dr. Bruton discovered agammaglobulinemia in 1952, has just grown exponentially and we now are up to at least 120 different syndromes that have been described over the past 53 years. The main point is that these conditions are usually recognized only when the person gets sick because outwardly these people don't look any different than a normal infant, child or adult. Then, the other thing to remember is that if you don't recognize it, and usually they are not recognized, then the problem is that you end up with permanent organ damage. Dr. Ochs has already touched on one of these. This would be central nervous system damage if one develops a persistent enteroviral infection. This is a slide that I use when I speak to the residents at our institution to try to help them think about these conditions. In the past it has been helpful to think about the types of infections that you have if you have a B cell defect, or a T cell defect, or phagocytic cell defect, or a complement deficiency because there have been certain characteristic organisms that have been associated with these. That way, one can then select the test that would be more appropriate for this. You notice also on this slide that I said the acquired deficiency diseases such as those that we, as physicians, cause when we give chemotherapy or immunosuppressive agents for infections that alter the immune system such as HIV. They are thought to be more common but for those in the bottom half of this slide we really don't know what the incidence or prevalence of these conditions is. They are thought to be more rare and it is for that reason that they are not screened for in usual screening tests. So, the way that these patients really are recognized is that they have increased susceptibility to infection. The caveat there is that this has changed recently because primary care physicians use antibodies liberally. We have had patients referred to us who have been to their primary care doctor every other week and received another antibiotic when they left, and they were not diagnosed until they were 5 or 6 years old because they were covered by the antibiotics. So, the classic presentation of immunodeficiency with meningitis, septicemia or osteomyelitis you probably don't see very much anymore because of the use of antibiotics. Since they do appear to be normal individuals on the outside, if they are taking an antibiotic and they don't have a serious infection the only way you would make this diagnosis would be if you had a high index of suspicion. So, this has been the classic way of thinking about these diseases. If you have a B cell defect, which would be the type of defect that IGIV would be most appropriate for, the organisms that people who have antibody deficiency syndromes are infected with most commonly are all the various strains of pneumococcus, staphylococci, H. influenzae, streptococcal organisms, mycoplasma organisms, and we have already talked about the enteroviruses and Giardia. The site of these infections usually is respiratory, although it could certainly be septicemia or meningitis if one were not on antibiotics. Whereas, for T cell defects you would think of things like CMV, EBV, the herpes family of viruses, and opportunistic organisms like PCP, and the characteristic of these infections would be that they would be severe and persistent. Whereas, people who have phagocytic cell defects would usually have problems with things that are on the skin or on mucosa, such as staphylococcal organisms on the skin or Pseudomonas. Now, Serratia marcescens used to be sort of the hallmark of a patient with chronic granulomatous disease but that is changing, as I will talk about in a minute. They also have problems with fungal infections. Of course, the hallmark of a patient with chronic granulomatous disease is that they have boils, and if you have another type of leukocyte deficiency such as ALD, then cellulitis would be the presentation. There are new syndromes that have been recognized, that I will talk about in a minute, that are characterized by mycobacterial infections and by salmonella infections. Then, of course, deficiencies of complement components. In the early components you would think of pneumococcal infections or staph. infections. Neisseria infections would characterize those that had defects in the late components. Of course, people with complement deficiencies often also have autoimmune diseases. This has been sort of the standard way of thinking about these different diseases, but now that we are understanding more about the molecular basis of these our thinking is changing, and I will talk about that in a minute. Let me go back to the end-organ damage. This is a boy who has Bruton's disease and you can see that he has right middle lobe disease. So, if this condition is not recognized and infections with either pneumococci or H. influenzae or Pseudomonas develop in these patients, then they are going to get bronchiectasis. There have been recent surveys of people who are adults who have Bruton agammaglobulinemia, for example, who are living now into their 30s, 40s or 50s. I guess the encouraging thing that has happened since IGIV has been on the scene is that there are fewer patients now with chronic lung disease. Somewhere between 20 and 30 percent of the adults who are surviving who have agammaglobulinemia do have chronic lung disease, and many of these die from this condition. The other thing to remember is that not only do they have lung disease but they have pan-sinusitis and, no matter how many sinus surgeries they have, they still are going to have pan-sinusitis. The role of rotating antibiotics, of course, is crucial in the management of this because since IGIV contains only IgG there is no way of replacing your mucosal immunity, and even if it contained IgA, Dr. Stiehm, in his study, showed years ago that there would be no way to get the IgA out on the mucous membrane surface. Hans has already talked about this so I am not going to dwell on this, but the reason that people who have agammaglobulinemia get these infections is that they don't have secretory IgA. So, when they have summer diarrhea and they develop echovirus infection, then the IgA is not there to prevent entry of this virus so then you have entry of the virus into the circulation, and then it crosses the blood-brain barrier. Another example of this, of course, is the live polio vaccine and many of the patients who have had these enteroviral infections have had this either from vaccine virus or from wild type polio virus. As Hans has already said, this condition still occurs. Fortunately, in a recent survey that was done by Jerry Winkelstein's group the incidence of this has dropped dramatically since IGIV was developed. But, as in the case of Dr. Stiehm and Dr. Ochs, we have now 7 or 8 patients at our institution who have CNS disease and we have tried to identify the cause of this, and in most of these cases we have not been able to tell what the etiology of the central nervous system infection is. The other end-organ damage, of course, is bronchiectasis. This is an adult patient with CVID who has bronchiectasis. Of course, this could be prevented by preventing the infection in the first place. Many of the patients with CVID--those of you who are not working in the field of primary immune deficiency may not appreciate the fact that the molecular defect in most cases of CVID is unknown, but there is a phenotype where there is lymphoid interstitial pneumonia or there is splenomegaly or this is lymph adenopathy and this seems to be triggered by infections, and we don't know which infections are the main triggers but they do have B lymphocytes and perhaps these B lymphocytes can proliferate even though they can't become plasma cells and make antibodies. This is just a CT showing the enormous spleen and adenopathy here in the peritoneum in a patient with CVID. Often, as a consequence of the splenomegaly, they will have penias, such as thrombocytopenia and anemia and neutropenia. Now that we are understanding what the molecular defect is in many of these conditions, we have to think about this in a different way rather than just thinking about B cell, T cell, phagocytic cell and complement. For example, in a recent review written by the group in France, whenever one has recurrent pneumococcal infections, obviously you would think of a B cell defect, but also those who have T cell defects because you need to have T cells to help the B cells. Of course, we know that defects of the early complement components can also lead to pneumococcal disease. Congenital asplenia is something to think about. But there are two new conditions that are characterized by pneumococcal disease. One of these is NEMO, nuclear factor kappa-B essential modulator, also known clinically as ectodermal dysplasia with immunodeficiency. IRAK4 deficiency is one of the new late innate immune defects that are characterized by pneumococcal disease. So, whenever a clinician sees a patient with recurrent pneumococcal infections, they have to think about a whole variety of different defects now that might be the basis for this. Another concept is if one has a patient with mycobacteria and salmonella infections, then you have to think about either of interferon gamma receptor defects, IL-12 deficiency or IL-12 receptor deficiency or STAT-1 deficiency. All of these have been characterized by selective susceptibility to these mycobacterial organisms or to salmonella. If you have a patient that has chronic Cryptosporidium infections or Pneumocystis carinii--these are not things that you would normally think of with an antibody deficiency but patients with X-linked hyper IgM which, as Hans has already pointed out, is really a T cell defect--have infections with these two types of organisms. Previously we used to think that Serratia marcescens was the thing that was going to alert you to the patient with CGD but now we are seeing all sorts of unusual infections in CGD patients, such as Trichosporon pullulans and penicillium infections; a lot of candida infections. This was reported recently by Dr. Bill Shearer's group in Texas. So, there are lots of organisms that seem to be colonizing these patients that we haven't known about in the past. Then the new threats, and I think Dr. Scott is going to be speaking about this in her talk, are the West Nile virus; smallpox or exposure to family members immunized with smallpox; anthrax. Then varicella is going to continue to be a problem. Because VZIG is no longer available there will have to be some new way of thinking about how to prevent this type of infection. So, what I want to talk about is how you get to the diagnosis without having infection. Just getting back to what I said in the beginning, the incidence and prevalence of primary immune deficiency diseases are really unknown. There are various estimates but basically we don't know. I suspect that there are a lot of people who die of pneumonia or meningitis or respiratory infections where the death certificate says the cause of death is pneumonia, but it is possible that these people probably had primary immune deficiency. There is no newborn screening, and even with the revised screening list that is possibly going to be coming out soon primary immune deficiency is still not on the list. If you live in South America or in the Middle East where you get BCG on day one of life, this is almost certain death for those who have defects in T cell function. As I have already said, because we live in an antibiotic era we don't have a classic presentation. A survey done by the Immune Deficiency Foundation, just a year or two ago, found that the average time from the time of the first infection to diagnosis was 9.7 years. These were the types of infections they had. They had--I can't read the top one up here--sinusitis, pneumonia, ear infections and bronchitis. These were the leading types of infections. Of course, this is what you would see if you were a primary care physician. You would see these types of infections and they are treated easily with antibiotics or supportive therapy. But when they occur repeatedly one should think about these conditions. So, the current status of this situation is that there is no screening for any genetic defect of the immune system at birth or at any time during life, anywhere in the world. This is a major problem, as I have said, where a lot of vaccines are administered on day one of life and if you have a defect in any kind of cell such as a T cell or monocyte, then generally these infants will die. The paradox is that the screening methods are available and could easily be implemented if screening for these defects were accepted as a standard of care. The obstacle is that they are considered to be so rare that screening for them would not be cost effective. Population surveys that have been done suggest that primary immune deficiency affects an estimated 50,000 persons in the U.S. and that these are at least as common as hemophilia, cystic fibrosis, Huntington's disease and phenylketonuria. However, we won't really know the true incidence until there is population screening. In the IDF survey it was determined that half of all the people who have primary immune deficiency were not diagnosed until they were adolescents or older. So, speaking of cost, the cost of late diagnosis is a heavy burden of disease on the patient and leads to early demise. The majority of patients report two or more hospitalizations before diagnosis. The cost of hospitalization of these patients far exceeds what it would cost to screen for the defect and to implement therapeutic or preventive measures. This is from the IDF survey. You can see in this pie diagram that more than half of these patients have been hospitalized more than two times in their life before the diagnosis was made. So, the only thing that really exists in terms of incidence data comes from blood banks where they are screening the donors and recipients because people who have IgA deficiency are often the ones who have unexplained transfusion reactions because of anti-IgA antibodies. A blood bank in Knoxville, Tennessee, where they screened something like 6,000 blood donors, found that one out of every 333 people who donated blood--and these are people who had to fill out a card saying they had no chronic infections, no chronic disease--one out of every 333 of these repeatedly normal donors had no IgA. IgA is the first cousin of common variable immunodeficiency. People who have IgA deficiency can eventually become common variables, but genetically it appears to be related very much to common variable. So, if you do the math and divide 333 into 280 million, it is pretty close to one million people in the U.S. who have IgA deficiency or some other form of primary immune deficiency. This would be just a guess. IgA deficiency could be screened for by measuring serum IgA on a heel stick done when the hemoglobin is first checked in infants 10-12 months of age. You could do the same thing when the child is getting ready to go to kindergarten or to first grade when the pre-school immunizations are given. You could also check again when they go to college. These would be times when you would have captive groups of patients that you could screen for this defect. If the IgA level is found to be low then, of course, you would measure the other immunoglobulins, and if they are found early enough IGIV can be started before organ damage occurs. The final comment about this is that there are existing genomic and proteomic methods that would make screening for all of these defects for which the molecular basis is known possible at birth. Turning now to another disease, and this is severe combined immune deficiency, these are 32 infants at our institution out of 144 that we have transplanted, who died after bone marrow transplantation. You can see that CMV, adenovirus, EBV, enterovirus--most all of these deaths were caused by viral agents that they were infected with when they presented. None of them died of graft versus host disease. So, again, if one could recognize these defects early enough, before infections occur, then obviously organ damage or death could be prevented. So, what is the prevalence and incidence of SCID? Well, it must be very low because it is uniformly fatal in infancy unless it is corrected by immune reconstitution. What is the incidence? Again, this is unknown. The literature says anywhere between 1/100,000 to 1/500,000 but I suspect it is much higher, probably at least as common as something that is screened for routinely which is phenylketonuria. Just to give you an example of an existing screening test that is available for this, if one did a white count and a manual differential on the cord blood you could determine the absolute lymphocyte count. Since SCID is characterized by an absence of T cells, that means you are missing 70 percent of your lymphocytes. So, all of these babies are going to be lymphopenic and that means that if you just recognize lymphopenia in the cord blood then the next step could be taken, which would be to do flow cytometry to see if T cells are absent. This is just to give you an idea about lymphocyte counts. Normally they are much higher during infancy than they are during older childhood and adulthood. Unfortunately, the time that most of the babies were referred to us presented was around 6 months. The lower limit of normal is 4,000. Many of these patients had had blood counts done repeatedly and they weren't recognized because people didn't realize that lymphocyte counts are normally much higher during infancy. This is just to show you the mean lymphocyte count for all the different types of SCID at our institution. Here is the lower limit of normal and you can see that all of these are low. There is one exception here where there were maternal T cells present that gave a very high lymphocyte count but, even so, it was still below the lower limit of normal. Just to show you how this could be applied, this is a pedigree that Dr. Jennifer Park had studied and found to be effective with X-linked SCID, and these are the carrier females. This mother was pregnant with twins and, for religious reasons, they did not want to do amniocentesis or chorionic villi sampling to find out whether or not the boy was affected because they were hoping that the little girl could be a donor for the little boy if the boy were affected. When she went into labor--they were delivered in our institution, you can see that a white count on the cord blood revealed that the little girl had a normal white count and a normal absolute lymphocyte count, but the little boy had a low white count and a low absolute lymphocyte count. We immediately did flow cytometry, and you can see that the boy had essentially no T cells or natural killer cells. So, this diagnosis could be made at birth and we were able, on the basis of this, to transplant this baby. You can see 6 months later that the little boy was as healthy as the little girl. It turned out she could not be the donor so we used the mother as the donor. So he is chimeric with his mother's T cells. This is just to show you, because we have seen a lot of SCID infants at our institution and even though we have given genetic counseling, often they conceive other children. So, we have had cord blood shipped to us for studies right at birth. We had 25 SCID infants that we had delivered at our institution but then these were siblings. These were 14 healthy newborns. You can see that there is a little bit of overlap in the absolute lymphocyte count. These are the normal controls and these are the SCIDs. But if you set the cut point at around 2,500 and then do flow cytometry you would be able to determine. Another way that you could do this would be to base it on the fact that there is absolutely no overlap at all in the T cell count for the SCIDs versus the normal infants. So, a test that is based on T cells would be something that would be even more reliable than just the lymphocyte count. So, as I have said, if you have found lymphopenia you would repeat this just to make sure it wasn't a lab error and then you would do flow cytometry. And, if there is an absence of T cells the diagnosis would be confirmed by T cell functional studies but the patient would be placed in isolation. You would have this information immediately at birth. Of course, if you made the diagnosis the condition could be treated without doing any pre-transplant chemotherapy, and many times this can be done as an outpatient and the cost of doing this as an outpatient is around $50,000 whereas, if the child gets sick and comes in at 6 months of age it can cost up to a million dollars for intensive care unit treatment. This is just a Kaplan-Meier to show you 39 SCIDs that we have been able to transplant in the first 3.5 months of life. You can see that we have only lost two, one from CMV and one from EBV. I am going to skip this and talk about this. One of the objections that we have to the suggestion that routine lymphocyte counts be done on cord blood is that the neonatologists prefer to do all their newborn screening on the Guthrie spot. The Guthrie spot is a filter paper test that was developed by Dr. Guthrie 30 years ago when he was interested in diagnosing phenylketonuria. It is 5 drops of blood on a piece of filter paper that is shipped off to the state lab. On these drops of blood you can do many, many different tests. Most of these are done by mass spectroscopy. However, it would require a DNA-based test to do a study that would detect T cells. This is just a cartoon showing you the basis of the test that is currently under consideration. Whenever the germ line configuration of the antigen receptor genes is changed by rearrangement of these genes there are pieces of DNA that are cut out, and these pieces of DNA are excised form a circle and they are called signal joint TRECs. So, a new T cell that has just recently rearranged its antigen receptor would have lots of these pieces of DNA that have just recently been cut out. Just to show you, we had blood stored on one of our SCID patients from pre-transplant and you can see that there are no TRECs present prior to bone marrow transplant. After bone marrow transplantation there are normal numbers of these pieces of DNA that had been excised. So, it is a very useful marker for T cells. Newborn infants characteristically have about 99 percent naive T cells so these naive T cells would be loaded with signal joint TRECs and, therefore, the Guthrie spot could be eluted and in a DNA-based assay one could analyze for this. This is just to show you all the ones that we had pre-transplant blood on, and you can see that there are only two that had any detectable TRECs at all. The lower limit of accuracy is around 100 here. Even placentally transferred maternal cells don't interfere with the TREC assay. Jennifer Park and her group have recently developed an assay that can be done on the Guthrie spot. By taking just two punches out of one of these Guthrie spots you can extract the DNA and you can quantify these TRECs and then this would be a measure of T cells. She was able to get from the State of Maryland the filter paper spots that had been shipped in on two babies that she later determined had X-linked SCID. So, she was able to retrieve these Guthrie cards and show that they did not have any TRECs, whereas these are various controls here that were all positive for TRECs. So, the TREC assay is a promising tool for large-scale newborn screening for SCID and future studies will determine whether or not this is going to be practical. Many sites don't have any DNA-based testing for newborns but I think this is all going to change in the future. So, if you don't have screening, then how else can you find these people before they develop infections? One of the things that the Immune Deficiency Foundation is trying to do is to develop clinical care guidelines. A clinical care guideline or practice guideline was defined by the Institute of Medicine as guidelines that are systematically developed statements to assist the practitioner and patient decisions about appropriate health care for specific clinical circumstances. These statements can also be used for quality improvement and payment policy making. So, last spring the Immune Deficiency Foundation had a committee that was charged with developing these guidelines and these are, hopefully, going to be released in the not too distant future. They are written so that patients and their families can understand them and call their physician's attention to the most appropriate testing and treatment that is currently available. Since there is no stronger advocate for the patient than the patient himself or the family of the patient, hopefully, by having this in a site where patients and families can read this, even if the physicians caring for the patient are not a hundred percent up to date on primary immune deficiency or the treatments, at least the parents will be able to notify these physicians about this. So, hopefully, this will help early detection and prevent organ damage. Thank you very much. Yes? PARTICIPANT: I think, if I understood correctly, you were saying that in Bruton's agamma you were still seeing the 30-40 percent increase in chronic lung disease at ages of around 30-40. Is that true? Did I cite you correctly? DR. BUCKLEY: Well, the surveys that have been done, they have been done on adults who are now, you know, between 20 and 40. IGIV, as Hans said, was not put on the market until 1981 so these people who are adults now would not necessarily have benefitted from IGIV throughout their lifetime. But there are still some cases that are occurring and, as Hans said, even in those who are getting IGIV so there are unknown infectious agents that appear to be affecting these patients. PARTICIPANT: The point that I wanted to make is actually a challenge to current immunological dogmas that IgA is so important for protection against mucosal immunity. I mean, there are two reasons why I think that should be challenged. One is that patients with selective IgG deficiency rarely get into trouble with infections. The other is that studies with vaccines have shown that if you get high enough IgG titers they actually diffuse across mucosal areas, and in the AIDS field papers have been published showing protection with only IgG in the vaginal mucosa protecting against AIDS. DR. BUCKLEY: Well, you are absolutely correct. The reason that people with selective IgA deficiency don't get this is that even if the enterovirus penetrates the gut mucosa and gets into the intravascular compartment they have circulating IgG that can deal with it. But the problem is that the gamma globulin preparations, as Hans said, didn't necessarily contain high enough titers against all of the different serotypes of the echoviruses. So, it is possible that a person can mount a very strong IgG response to that specific ECHO type and can deal with it. We monitored actually patients with IgA deficiency for these enteroviral infections and found that a number of them did have them in their GI tract but we never found them in their bloodstream or in their CNS. So, I think you are right that IgG can deal with it and I think that accounts for why we are not seeing as much of it as we did previously. PARTICIPANT: Can I ask you a question? DR. BUCKLEY: Yes. PARTICIPANT: When you were talking about screening [not at microphone; inaudible]... a subset of individuals with IgG deficiency who also have [inaudible]... would you give an estimate or guesstimate as to [inaudible]... IgA deficiency [inaudible]... IgG replacement therapy. How many IgA deficient individuals do you think you would have to screen that way in order to identify one who had an IgG deficiency? DR. BUCKLEY: Well, I really don't know because, you know, it all gets back to the fact that we don't know what the incidence or prevalence of these defects is since no one ever tests for them. But I know that for families that I have evaluated where there has been IgA deficiency there has been, like, a sibling who had common variable. I have seen IgA deficients convert into common variables. So, even if you just detected IgA deficiency it would be worthwhile doing because these people do have health problems. They have autoimmune diseases. They have increased risk for cancer. There is a whole lot of other reasons to screen for that, let alone the fact that they would be at risk for transfusion reactions if they were to be given a blood product that contained IgA. Thank you. Surrogate Markers for IGIV Licensure DR. STIEHM: It is a pleasure to be here. Whenever I give a talk early in the morning in a different time zone I am reminded about my experience in Amarillo, Texas. I was asked to give a talk at eight o'clock in the morning and I get there, and there is an auditorium about this size but there is only one person in the audience. I thought, oh my God, I must have the wrong time but I looked and it was the right time; and I thought it might be the wrong date and I checked my calendar and it was the right date. So, I gave my talk and the man applauded, asked me a good question and then as I was leaving the podium he said, "where are you going?" And, I said "I am going back to Los Angeles." He said, "you can't go; I am the second speaker." [Laughter] What I am going to speak about this morning is a possible alternative to our current method of allowing new IGIVs on the market. Currently, if you want to develop a new IGIV and want to release it you have to round up around 40 antibody-deficient patients that have profound deficiency and give that individual IGIV for 3 or 4 weeks for about a year, 400-600 mg/kg/month, and compare the frequency of infections with the prior IGIV. Very often pharmacokinetic studies are done at the fifth or seventh month on a subset of these patients. Immunoglobulin levels are measured at several points, and you record serious infections, and the FDA now requires one or less per year, other infections, fevers, antibiotics, hospital days, absences from school or work and possibly physician visits. So, is there a need for surrogate markers for IGIV testing? Well, first of all, new products are entering the market. Secondly, patients with XLA and common variable immunodeficiency, the ideal patients to test this new product, are fairly rare and they may not want to participate in a new IGIV trial and have all these blood tests done, etc. And, the current trials may not pick up an illness, such as varicella which these patients are susceptible to, even though a crucial antibody is missing because in that year they simply are not exposed to that. So, a proposed alternative for IGIV licensure--well, first of all, the IGIV must have a good safety profile. It can't be infected with pathogens, and the side effects must be acceptable, as people have talked about and as we will talk about more. Secondly, the IGIV should have a good half-life, around 25 days, and I suggest that maybe that is even not necessary because the trough level will parallel the IGIV metabolism. The IGIV must provide adequate trough levels of IgG and IgG subclasses. So, I propose that maybe surrogate markers could be acceptable. Well, what are the surrogate markers that have been suggested for IGIV? First of all, IgG and IgG subclass levels; perhaps IgG or antibody for pharmacokinetics measuring the rate of disappearance of the gamma globulin or selective antibody. People have used sinus or chest x-rays or CT scans, possibly pulmonary function tests, possibly inflammatory markers as a way to assess frequency of infection, and my suggestion is maybe antibody titers would be the ideal way to do this. First of all, what about IgG level as a surrogate marker? Certainly the IgG level, the trough level is a very strong indicator of the amount of clinical benefit. It has been well documented that moderate dose IGIV of 400-600 mg/kg is superior to a lower dose, 100-200 mg/kg, in terms of decreased antibiotic usage; decreased days of fever and hospitalization; improved pulmonary function; weight gain; clearance of sinusitis. So, we have this important surrogate marker of simply the IgG level. A recent paper suggested maybe even higher doses of IGIV may be better than what we now sort of regard as standard dose. This is a study out of The Netherlands, showing that for patients who received higher doses of 600-800 mg/kg, essentially double of what they were previously getting of 300-400 mg/kg, the number of infections decreased. The frequency and duration of illness decreased. Of course, the IgG trough levels increased dramatically. So, here you are comparing 660 mg/kg to 940 mg/kg and there was clinical benefit with the possibility that, as people have brought out, maybe you are getting more gamma globulin into the secretions by giving these higher doses. Of course, they are much higher. This is the trough level; the lowest level. Another concept is that there have not been demonstrated any clinical differences if the trough levels are comparable between different gamma globulins. In fact, Zuhrie et al. studied 3 different products in a comparison and could find no clinical differences of different IgGs. Schiff et al. also found in a study that there were no clinical differences in 4 different commercial products. More recently, Roifman et al., and we will probably hear more about this from Dr. Gelfand, studied a Bayer IGIV and found an incidence of acute sinusitis, 16/73 in an old product versus 7/73 in a new product, and the claim was that this showed that this was a better product than their previous product. However, there were no differences in 8 other infections and no antibody titers were done. So, I think this study as to be taken with a grain of salt. What do we know about IgG as a surrogate marker? Well, an optimal level is over 500 mg/dL. This level, as Dr. Ochs points out, is dependent on the dose, the frequency and the route, and I think that everyone agrees that milligram per milligram the subcutaneous will give you a higher trough level than does IGIV. Now, IgG level is a very good correlate of IgG catabolism so maybe these pharmacokinetic studies are not necessary, and probably repeated subclass determinations are not necessary either. We know that the IgG subclasses do differ in metabolism and biologic activities, and more recent studies show that IGIV usually has a normal distribution of IgG1, IgG2, and IgG3, and IgG4 doesn't really make a lot of difference in my opinion, and with most of the studies showing that the IgG subclass trough levels are similar to IgG in the proportion that they are in the serum. Dr. Ochs and his colleagues used pharmacokinetics as a surrogate marker, and he showed, like most other studies, that if you measure the half-life of patients with antibody deficiency they actually have a more prolonged half-life and that the subclass half-lives were also prolonged. The study shows, like in other studies, that IgG has a shorter half-life than the other, but it is pretty good in antibody deficiency because they do have longer half-lives. Morell et al. also used the disappearance of antibody to measure the pharmacokinetics of IgG, and again showed that in antibody-deficient patients the half-life of antibody is prolonged for 5 different antigens that are described here--hepatitis B, cytomegalovirus, tetanus toxoid, pneumococcus and H. flu. So, what do I think of IgG pharmacokinetics? It is easily done. You can do it using antibody or IgG levels after 6 or more infusions to get rid of the old gamma globulin and that may be unnecessary because trough levels give you similar information. Well, how about using pulmonary function tests as a surrogate marker? By the way, your handout has omitted four of the slides that I am giving right now. Simple pulmonary function tests, like FEV-1 or FVC, are readily available--no good in children under six, of course. Pulmonary function tests are okay in adults if they have a lot of lung disease but for most of the patients I don't think that they are a very good surrogate. Chest or sinus x-rays might be a useful surrogate but, again, these often don't change dramatically, particularly in the short period of time you are testing for a new IgG. What about the use of acute phase reactants? Cunningham-Rndles has some data using C-reactive protein, finding that it is a useful surrogate of frequency of infection. Several years ago I did a research survey of mostly infectious disease literature and came up with the 4 that I thought were the most useful surrogate markers for chronic infection, and these include a C-reactive protein, a procalcitonin, an IL-8 and mannose-binding lectin. There have been a number of studies to suggest that a combination of these 4 might be better than any single one directly. Others, such as the sed. rate, C3, IL-3, lactoferrin and a number of other ones have been proposed but I think that some combination of surrogate markers might be useful if you are drawing, say, a blood sample. Finally, what about antibody titers as a surrogate marker? I think these are probably the best. You have to consider which antibody should be measured; how should they be measured; what is the protective level; how often should they be measured; and are serologic assays equivalent to functional antibodies like hemagglutination titer versus a neutralization titer for a virus? You have to consider what antibodies are in a donor pool; what are the most important illnesses that these patients are likely to get; what are the most important pathogens; and what are the less common antibodies that should be measured, protecting against smallpox. Well, as Dr. Buckley has alluded to, we know that patients with antibody deficiency are susceptible to a number of bacterial and viruses, as well as mycoplasma, ureaplasma, cryptosporidia, pneumocystis occasionally, and bacteria as listed above, particularly S. pneumoniae and H. influenzae. Enteroviruses, of course, are very important as we mentioned. Well, if we look at what infections are occurring in these patients that are target patients, pneumonia, ear, nose and throat infections, gastrointestinal infections, etc. are common but other, less common infections, serious infections like meningitis, septicemia and osteomyelitis do occur and must be considered. What is the cause of death of patients with XLA and common variable? Pulmonary infections are way much higher, particularly since we no longer seem to have as much trouble with viral infection--echovirus and polio virus were high in the past but are less frequent now; liver disease; hepatitis C; lymphoma; gastrointestinal disease; sepsis and miscellaneous. When we consider what antibodies we should measure, first of all we ought to consider what do we immunize normal children for. We now have a whole laundry list of vaccines that are given routinely--diphtheria-pertussis-tetanus, measles, mumps, rubella, Hemophilus influenzae type B, hepatitis B, polio, varicella zoster and pneumococcus. Do we immunize high risk children? And these are, indeed, high risk patients. These include Pneumovax which picks up the other serotypes not present in Prevnar, meningococcus, RSV infection we don't immunize; we give them passive antibody; influenza, hepatitis A and then we have to consider what less common diseases do immunodeficiency patients get, such as Parvovirus, salmonella, Giardia, other respiratory viruses, hepatitis C, Cryptosporidia and then, of course, our sponsor here wants us to have a immunoglobulin that has adequate antibody titers to hepatitis B, diphtheria, measles, tetanus and polio. These are the statutes of the FDA. So, what are the choices of antibodies? Well, my list includes what is common in primary immune deficiency and these are the pneumococcal titers and the H. infuenzae titers. What do these patients need prophylaxis against? Measles, tetanus, diphtheria, polio, hepatitis B, hepatitis A and chicken pox. What is ubiquitous and unavoidable? Cytomegalovirus. Well, do the antibodies vary from lot to lot? They certainly do for less common organisms. This is just one recent example of a study comparing antibodies lot to lot for E. coli, Staph. aureus, Staph. epidermidis and enterococci. The titers in Sandoglobulin versus Gamimune were strikingly different. Even more important, Weisman in 1994 looked at both manufacturers and different lots from the same manufacturers and there were tremendous differences for E. coli, pneumococci, H. influenzae, Staph. epidermidis and group B streptococci. So, even product to product it is going to vary tremendously probably depending on the plasma pools that they are derived from. So, my conclusion is that there are significant differences in antibody titers to IGIV, probably less common for major pathogens like pneumococcus and Hemophilus, and these differences are particularly crucial if you are studying less common disease such as nosocomial infections in newborns, CNS enteroviral infections, RSV or EBV virus. So, the antibody titers that I would test for if I was going to do surrogate testing for a new IGIV product would include H. influenzae; pneumococcus, 5 serotypes; diphtheria; tetanus; hepatitis B; measles, varicella zoster and cytomegalovirus. Polio is required in the product and wouldn't be used as a measure of efficacy because it is a very difficult antibody to measure and you get such low titers. First of all, let's just discuss these in brief, what we are talking about. For pneumococcus, it is the most common organism causing respiratory infections in immunodeficient subjects. We know what the protective titer is and we know that IGIV donors usually have protective titers to most of these, particularly when you pool them. There was a product on the market several years ago called bacterial polysaccharide immune globulin. The plasma pool was derived from individuals who were given immunization to Hemophilus, pneumococcus and meningococcus and these had extremely high titers. But in 5 percent IGIV there is a fairly good titer. You can read the numbers in the handout. Following IGIV you get adequate trough levels. Dr. Scott has a paper that is going to be published that details these titers in more detail. I think it is important to assess several serotypes by ELISA that are absorbed with cell well protein and all of these are present in Prevnar. One important consideration is should opsonophagocytic titers be done, and I would think that for the initial studies you should probably do both to make sure that these do correlate. Then you might want to consider other serotypes that are not present in Prevnar but are present in Pneumovax. Hemophilus influenzae is a second most common organism causing respiratory infections in PID. Again, 5 percent IGIV has adequate amounts of it. The post-immunization titers are greater than 300 ng which is well beyond the protective levels, and there is sufficient antibody in the trough levels following 400 mg/kg of IGIV that gives you protective titers. For diphtheria it is required by the FDA--a rare epidemi--and, again protective titers are present although no one has ever measured the trough level of this particular antigen. Every one needs protection against tetanus. Indeed, there was a study to show that you could use IGIV instead of tetanus immune globulin to provide adequate titers of tetanus antibody following injury that exposes you to tetanus. C. tetanus I think would be a very good antibody to have in this pool. Measles, again, is one of these ubiquitous organisms. FDA requires that it be present in IGIV. There are a number of assays available. The neutralization titer is sort of the hallmark of protection. And, we also know that IGIV inhibits responsiveness to measles vaccine following a large dose. For example a child with Kawasaki disease can't be immunized for 4 months following a large dose of IGIV. There have been limited studies of measles titers following an IGIV infusion and titers seem to be protective. Chicken pox is ubiquitous and this is a serious disease in immunodeficient children. So, it is present in IGIV, particularly present in varicella zoster immune globulin which is used for immediate prophylaxis. There is a study to show that you can protect against varicella with 300 mg/kg, and rarely do patients on IGIV get varicella and, if they do, it is usually a very mild course. Hepatitis B is also required by the FDA. These kids are exposed to multiple needles, procedures, personnel and they seem to be unusually susceptible to hepatitis B and IGIV has adequate levels of hepatitis B and, of course, hepatitis B immune globulin has extremely high titers. Finally, cytomegalovirus would be a good one to test for. It is an ubiquitous infection. It is present in blood products that these patients are exposed to and it often will cause severe infection in primary immune deficiency. We don't know what the protective levels are. We do know that IGIV contains cytomegalovirus antibody and cytomegalovirus immune globulin contains titers that are 2 or 4 times higher. Some of the other organisms that might be considered for antibody titers would be Staph. aureus., Staph. epidermidis, E. coli, pseudomonas, HSV1 and 2, Coxsackie, echovirus, parvovirus and possibly others. Other surrogate markers that have been used for IGIV for other indications--for ITP, of course, the platelet response and the duration; for Kawasaki disease, prevention of aneurysms, possibly inhibition of cytokine production and neutralization of super-antigens; for newborns it is particularly useful to have antibodies to Staph. epidermidis or Group B strep. and possibly opsonophagocytic index might be more important. For transplant patients cytomegalovirus is a crucial antibody and possibly the inhibition of a mixed leukocyte culture. So, my proposal for an IGIV protocol based on surrogate markers is to give IGIV for a limited number of patients; of course, do safety and infection monitoring; and measure IgG, IgG subclasses and antibody titers 4 times, at pre, 4, 8 and 12 months. You might want to do pharmacokinetics but I don't think that is really necessary; then have these sera available for antibody titers to other organisms. The patient would be monitored pre and at several points during the trial, and you should also assess past and current IGIVs and measure these levels in normal adults, young children and older children, and possibly immunodeficient patients pre-treatment and after treatment as good controls. So, are all IGIVs significant? Well, there are significant differences in side effects. Antibody titers are usually similar for major pathogens; variable for less common pathogens and probably for functional assays. The half-life has minor differences. The achieved IgG trough levels have minor differences. Achieved IgG subclass levels have minor differences and the clinical efficacy has minimal differences. In summary, I think we want a trough level of greater than 500 mg/dL. We want subclasses distribution similar to the IGIV administered; antibodies titers to the 12 antibodies that I mentioned. Pulmonary function tests and acute phase reactants may be done but I don't think are crucial. And, pharmacokinetics, functional antibody assays, x-rays and other antibody titers might be considered. Thank you. DR. SCOTT: Can we ask speakers to identify themselves? DR. PIERCE: Yes, this is Dr. Pierce from the FDA. What can you tell us about the statistics that would help us to better understand where we are in validation of using and relying on the trough levels, realizing that you also recommended other things? For example, in the study that you mentioned that was a randomized study comparing a higher range of doses versus a lower range of doses I believe that the trough levels seen in the higher range dose group were much higher than 500 mg/dL. Also, in running through the list of several authors who had looked for differences between different IGIV products in their studies, could you tell us something about the numbers there? What was the biggest difference that could have been missed between any two IGIV products given the sample size that they examined? DR. STIEHM: Yes, I looked long and hard for head-to-head studies and they are really very minimal. I think I found three and they were a very limited number and certainly didn't divide into type of infection. But my overall karma is that there have not been significant differences from one brand to another unless the trough level is different. I think that there is considerable evidence that trough levels are probably the best surrogate marker for infection. DR. PIERCE: I guess my perspective would be that if it is still an open question as to whether there are differences in efficacy for prophylaxis of infections between different IGIV products that would be given in a dose that produced an identical trough level, then it would also follow that the reliance on the trough level--that the jury may still be out in that regard. DR. STIEHM: Yes, I agree with that and that is why I think antibody titers in addition to trough levels would be important to measure. IGIV for Primary Immune Deficiency: Antibodies Against Potentially Problematic Pathogens DR. SCOTT: Just a general comment before I start, and that is I think we are in the position now of asking whether we need to and should deconstruct IGIVs with regard to antibody specificities, and whether there is a way of knowing whether right now the products are optimally protective. I would point out that our impression at any rate is that IGIVs are studied in a subset of selected patients for the efficacy trials and, of course, that might not relate entirely to the population in the field that needs to receive IGIV for prophylaxis against infections. But I think it would be useful to figure out if it is possible to learn if the IGIVs are optimally protective against the common pathogens and the less common pathogens, and one question I will have is how could we find that out? And, if we did find out that there was a need for improvement I think that would be very useful for the patients. We are a little behind time so I will try to go through this in a moderate hurry. Thank you, Dr. Stiehm for your talk. It was very helpful and related to what I am going to talk about next. Just as background, as was mentioned--all Ig products-- ctually we don't have anything in the Code of Federal Regulations for IGIV products--should have a minimum potency against diphtheria, measles and polio. Now, why did this occur or why was this regulation made? It was made in 1973. There may have been some different epidemiological considerations and also testing considerations but this was intended to be a surrogate for product integrity and effectiveness, these tests. They are meant to assure a certain amount of lot-to-lot consistency of immune globulins and, by the way, they should all be, according to the Code of Federal Regulations, functional assays. There are lots of reasons to measure other antibody specificities so that we can define the scope of product activity against the relevant pathogens that have just been discussed that commonly infect people with primary immune deficiency. They could also be used to address predictors of efficacy and they are important in viral safety in two fashions. One is that, of course, they have neutralization potential, particularly against non-enveloped viruses but also envelope viruses that may inadvertently contaminate the starting plasma pool. In addition, at FDA and elsewhere it has been shown that during manufacturing antibody complexed viruses are partitioned away from the IGIV product. So, they are also important for that reason, for example, for hepatitis B. Understanding the other antibody specificities may help us at least estimate the likelihood of protection for primary immune deficiency patients against emerging pathogens. That is a subset of things that I would like to talk about. IGIV is probably constantly in evolution with respect to its antibody specificity for the less common pathogens for which we are not vaccinated. So, donor epidemiology and manufacturing I think are the two main drivers of what antibody specificities you have. Donor epidemiology is based on disease exposures and vaccination status. I would point out that in the past decade at least, and longer, there has been a change where natural immunity is less often induced because people have been vaccinated against certain pathogens, and the question is outstanding whether these natural, so-called, antibodies from people who have had natural infection have a higher avidity or are more long-lasting, or present in higher titers. Of course, the people with these immunities to the natural infection are now getting older and their antibody titers may wane. But it has been shown for measles, at any rate, that the antibody titers from a naturally acquired infection are higher than from a vaccine. Manufacturing methods can affect the antibody's repertoire, particularly manufacturing methods that eliminate or damage IgG3 which is a more sensitive subclass to various chemical and enzymatic treatments. It can affect titers, for example, to measles antibody and others that have some IgG3 subclass represented. I am just going to briefly go through all of these. We have pathogens with a changing epidemiology because of vaccination. An emerging pathogen of interest is West Nile virus. We also have some that are introduced, at least these would be pathogens with certain immune deficiencies by the counterterrorism efforts and particularly people with primary immune deficiency, and the Immune Deficiency Foundation called our attention to this. We were worried when we had a mass smallpox vaccination potential and when the first responders were vaccinated that they would accidentally acquire vaccinia infection from vaccinated people and then become ill. Nearly everybody here has antibodies to measles and there are very few cases per year. There haven't been any epidemics since 1997 and, therefore, there is no boosting of the population of people who have already had measles or been vaccinated. Of course, the proportion of donors to plasma pools born after 1957 is increasing so now we have more vaccinated people than we do people who had natural infection. Because vaccination results in lower levels of probably high affinity antibody to measles and certainly lower levels, there has been a concern that measles antibodies will decline in immune globulins. We would like to understand whether this is the case. Judy Beeler, in her lab at the Office of Vaccines, in collaboration with M-Y Yu and myself and some others and Suzanne Audet, have done some measurements on IGIV products for measles neutralizing antibodies, and these are from 166 lots from the years 1998-2003. These are actually subsetted into years but I didn't have time to show you that. Basically the point I want to make from this slide is that here you are looking at the geometric mean titers so the higher the numbers you have here, the higher the neutralization activity, and these are just various different products that were looked at and this is the source of the plasma pool. What you will notice is that most of the products are quite similar but, interestingly, these two products are made using the identical manufacturing procedures and what you see here is that the source plasma lots tend to result in lower titers than the recovered plasma lots. This is actually true now, we know, for two different products where the recovered plasma still yields higher titers of measles neutralizing antibodies. In addition, we have a product here with a somewhat lower titer and this we think is probably a result of the manufacturing methodology. Varicella zoster is also a concern because people with immune deficiency are susceptible to severe disease. It is generally known that the supply of varicella zoster immune globulin may be severely limited due to non-manufacture in the future of this product at least by the current manufacturer. There is a protective titer that has been defined by vaccine studies. Of course, that is in normal people. But the question was already brought up whether or not IGIV itself, any IGIV, can decrease the varicella attack rate or the severity in primary immune deficiency patients. There aren't any really systematic studies but there are some case series starting with immune serum globulin looked at in the 1960s to 1970s, followed by more recent studies which show that IGIV may have provided partial protection as did VZIG to exposed pediatric oncology patients, so kids that were exposed to smallpox and were immune compromised. There is one letter to the editor by Ferdman, in Pediatric Infectious Diseases in 2000, where he had three people with primary immune deficiency that developed varicella while on regular IGIV treatment. These subjects have received IGIV 7, 11 and 30 days prior to the onset of their varicella infection. So, this tells us at least--and we don't know what IGIV it is and I am not really sure it matters--but this tells us that it is not necessarily protective. Now, none of these children got severe disease. That was the good news. But we don't have a handle on those titers and we don't really know what would be protective. But if VZIG is in short supply I think t is important for us to have an idea where the patients will stand that are receiving regular immune globulin. In collaboration with CDC, we started a research project. We have requested from our manufacturers 5 lots of each product and the testing for anti-varicella antibodies is being done by Scott Schmidt who is at CDC. He will be comparing these IGIVs directly to VZIG. The purpose of this is not to make a claim. Rather, it is to estimate the likelihood of protection for patients with the current IGIVs. We would also like to monitor trends in the products as the varicella vaccinated donors continue to predominate over the naturally infected donors over time. We are also interested in looking at antibodies to West Nile virus and IGIV. Certainly people, especially with these mixed immune deficiencies, are likely to be susceptible to severe disease if they contract West Nile virus. Now, IgG titers in donors were reported by Mike Busch at the BPAC last October and he mentioned that in epidemic areas up to 5.3 percent of donors might have IgG titers against West Nile virus. So, we would like to know whether or not there are yet any detectable anti-West Nile virus antibody levels in IGIV because anti-West Nile virus antibodies, at least in animal models, can protect against disease and, of course, we have concerns about the immune deficiency patients. So, a project that we would like to start is to assess the emergence of these antibodies in IGIV, the lag time between epidemics and emergence of antibody and whether there are regional differences, and the relationships of titers to geographic collection practices. Finally, I am just going to end with smallpox vaccination. The vaccination for first responders was announced. Half a million were supposed to be vaccinated. In fact, far fewer volunteered to be vaccinated but people with primary immune deficiency and the IDF are concerned about the potential for accidental infection, as I mentioned, and we wanted to find out whether IGIVs contain anti-vaccinia antibodies. We had an IGIV bank of products collected in about 2000 to 2002, and we sent these off blinded to a number of different labs to look at neutralizing antibody to vaccinia in the products. So, here you see the different products. These two are vaccinia immune globulins. This is the amount of antibody needed to neutralize 50 percent of the virus in culture. Basically, the lower you are here the better your titers because you need less to neutralize. What you can see is that all of the products had antibody to vaccinia and some of them might have had a little bit more than others. We also studied some of these in vivo. We just selected lots with sort of medium range titers and found that, indeed, when co-injected with vaccinia into SCID mice they could increase the survival time, which I think is a useful parameter to look at, SCID mice being the very worst case for susceptibility to infection. Finally, I want to end with a question. Is there a role for an IGIV repository for research purposes so that we can look at the antibody titers to different pathogens over time? This would enable us to monitor trends in emerging pathogen antibodies, and it could enable us to at least estimate the potential amount of protection or lack of protection in these products against new pathogens. The purpose would not be to compare products though. If such a repository were to exist, I would suggest that it would be comprised of 5 random lots of each product; that the aliquots would be coded and frozen; and that yearly deposits would be made into the bank by the different manufacturers. One important question I think, especially important to the providers of this material, would be who can request these for testing. I think certainly FDA would be among those and CDC as well, but also outside investigators and industry might be interested and that would have to be discussed with the people who are supplying the material. Should the samples be blinded when they are sent out? I think yes. If so, when or should they ever be unblinded? For FDA research, we would look at the antibody levels to common and emerging pathogens. We would code published information and, of course, any such program would be voluntary. There wouldn't be a requirement to submit these samples and that would be different from the lot release requirement. I think that is all I have to say here. Thank you for your attention. PARTICIPANT: The only question about the lot repository is that if you don't capture epidemiological data on the donors where you got the plasma from geographically you basically have a blind set of samples and it doesn't really give you the ability to tie it to particular kinds of plasma. I think you have to ask the manufacturers to put more information into anything they would deposit. DR. SCOTT: Well, there are two ways of looking at that. One is that more information would be very useful in that respect, especially where we see differences within a product, the lot-to-lot variation. On the other hand, people who are being treated with IGIV, of course, are receiving whatever lot happens to come to the pharmacy so in a way there is an advantage to looking at the spread. I wouldn't see any harm in collecting the information but we wouldn't want to target it to a certain epidemiology. An exception, of course, would be West Nile but that is a different matter and we are asking for samples separately anyway. Now to Dr. Schiff. I know we are running over time but I think in about 10 or 15 minutes we will take our break. Thanks. Are we Adequately Replacing Antibody in Patients with Primary Immunodeficiency? DR. SCHIFF: Thank you, Dr. Scott. Thank you for giving me the opportunity to talk to this group. It is an interesting position to be following my three mentors, Dr. Buckley, Dr. Stiehm and Dr. Ochs. My talk is going to overlap somewhat with Dr. Stiehm's because we didn't really talk about these ahead of time, but I am going to take a little bit different approach to this, that is, not so much what can we do to study new products but to look what we are doing now and are we achieving what we are trying to. The reason that I have had a concern about this for a long time, and Dr. Scott also alluded to this somewhat, is that IGIV is made from normal healthy donors. Healthy donors basically don't have very high levels of antibody. The whole idea is that we have low levels of antibody and then we get infected and you start making more of the antibody that you need. We have seen this in several patients who, when they got infected, their specific antibody disappeared very rapidly. So, when you have an immune deficiency patient, they don't have that opportunity and so what we are giving them is really sort of a low level protective antibody that healthy people have. Again as Dr. Scott alluded to, antibody titers may be falling. We have the situation of vaccine versus natural infection and I think she covered that pretty well. I am concerned about things like H. flu because, again, we are looking at immunized donors, adults that are now having children that are immunized who are not getting the high levels of exposure that they did before and with a lot of these diseases like H. flu we are not seeing as much. But there are also other reasons why. We have eliminated some of the high risk donors and I think we saw a fall in CMV titers after we eliminated some of the more high risk donors. We eliminated them because of the risk of HIV and hepatitis but, on the other hand, they also tended to have very high titers to a lot of other things that they were infected to. One of the things I am going to point out and go into some more detail on the data that Dr. Stiehm talked about is that high levels of antibody may be needed for chronic infections, and we know very little about the antibody titers during clinical studies. We have really focused pretty much on IgG levels, and I think that is important, but we really haven't focused very much on specific antibody. One of the things that we have found is that the recommended doses of IgG have increased over the years so what is that really trying to tell us? So, I will try to get into that a little bit. Then, also for the IGIV preparations, are the in vitro titers relevant? We often use different assays than we use for evaluating patients. If we are evaluating a patient to see whether they are adequately protected we really should be using the same assay, but also is there a difference measuring isolated IgG versus the antibody in plasma where that is the way the tests have been standardized? For many things we don't know the protective levels of antibody. Dr. Stiehm showed you a few where we do, but there are also a lot where we don't and are those the ones that are really causing problems now that we have solved sort of the easier problems? So, basically, are the current doses adequate? And, you know, my question is was anybody else besides me concerned? I think the fact that people in this room are here is testimony that we do have questions about this. To become somewhat biblical, in the beginning there was intramuscular gamma globulin, and immunologists looked down and said it was good. So, we were very pleased with ourselves in the 1950s, the morning and the evening of the first day. And, indeed, it was. I mean a lot of infections, sepsis, meningitis and severe pneumonias clearly decreased, so much so that in 1957--the chronic infections were less protected but in 1957 when the British Medical Research Council wanted to do a study they determined that to do a placebo-controlled trial was unethical. So, we have never really had a placebo-controlled trial of gamma globulin and, of course, I don't think we ever will. But they did look at some doses and they suggested that 100 mg/kg/month was a little better than 25 or 50 but patients weren't very well characterized back in 1957. We didn't even really know about B and T cells at that point, and 100 mg/kg/month was pretty much the upper limit that we could give to patients intramuscularly. Most people are just not tough enough to take more than that. So, we started off with 100 mg/kg/month and the first IGIV studies used that same dose. You heard, you know, that we achieved about the same levels and we got about the same results. In fact, there were a few more upper respiratory infections in the intravenous compared with the intramuscular but we thought that was due to the fact that for intravenous they came back every month and for intramuscular they came back every 6 months and they weren't keeping very good diaries and they just didn't remember how many minor infections they had. So basically they were equivalent. The first trial of high dose IGIV comparing 400 to 100 didn't show any significant difference, but the patients weren't carefully selected and, again, this was done quite a while ago. But Dr. Pirofsky did one study using 500 versus 150 and he found that if you really focused on the chronic infections there was a difference in the higher dose. There are some studies that Dr. Stiehm alluded to, and I will just show a little bit more data on that. This was a study published by Roifman and Gelfand in 1987, in Toronto, and the study design in this was 12 patients with hypogammaglobulinemia. All had chronic lung disease with pulmonary functions that were more than 25 percent below predicted. It was a 12-month crossover study, 200 versus 600, but there was no run-in or washout period. So, that meant that in the crossover there is a period of time when their IgG levels were changing but that was still part of the efficacy period. They evaluated the incidence of infection; measured levels and measured pulmonary functions. The thing that I wanted to focus in on is that if you just looked at the dose of gamma globulin being given you couldn't see very much difference, but if you focused in on the IgG levels you saw that if the patient had IgG levels less than 500 they had 23 upper respiratory infections versus 10 if it was over 500; 11 cases of pneumonia versus 3. So, the total was 47 infections versus 15 looking at the IgG levels alone. They also looked at the pulmonary function tests and, again, you can see that the pulmonary functions at the high dose--in the closed circles and squares, either FVC or FEV-1--when they were on the high dose were better and they declined when they were crossed over to low. That is true for the FEV-1 as well. You can see the opposite was true when they were on the lower dose, they improved after going to the higher dose. So, again, that is good evidence that the higher dose was better than the lower dose. This is another study that you saw some data on from Dr. Stiehm. This study was 9 months on the higher dose versus standard, and I will show you the doses in a minute; a 3-month washout phase and then crossed over. In this case we did have a washout period between the two. Again, the standard dose was 300 for adults and 400 for children. We have always done things on a kilogram basis and maybe for children we really should be thinking about doing this on a meter squared basis to try to correct for that. But they liked to give the children a little bit higher dose. Then for the higher dose they doubled it to 600 mg/kg or 800 mg/kg and I think, for the most part, all these patients were treated once every 4 weeks. But you can see again that on the standard dose the infection rate per patient was 3.5 or 2.5, with infection duration of 33 days versus 21 days. These are both significant. The days to first infection 82 versus 123. And you can see, as Dr. Pierce alluded to before, in this case at the higher dose they are getting levels of 9.4, very high. I remember when we first started treating patients we thought if we could get levels of 200 mg/kg at least for those who had some baseline IgG that we were doing well. Now we are starting to talk about levels of 9.4, which is getting up close to what the normal levels are in healthy adults. I am just going to go very quickly through their recommendations. They found that the high dose was better, but they still recommended that you start with the standard dose because some patients will do well. And, as Dr. Buckley alluded to, if you start patients when they are healthy and they don't have chronic disease they often will do better at lower doses and it is after they get chronic disease that it seems that we need the higher dose. Then, those patients that had persistent infections, more severe infections per year, then you increase. I am not sure that we should, you know, wait that long but the idea is that if the patients are doing well at low dose they don't have to go to higher doses. But many patients with chronic infections are going to need higher doses and maybe we really need to be achieving the levels of healthy adults. Then in a study that I guess Dr. Gelfand may talk about later comparing Gamunex to Gamimune, they also looked at the data comparing low and high doses of IgG, those over 400 versus less, and you can see that both for Gamunex and to a lesser extent for Gamimune there was a difference, but even more importantly, when you look at it in terms of trough IgG levels for both Gamimune and Gamunex you can see that there was a decrease, a progressive decrease in the incidence of validated infections. So, if you look at the total here--20 versus 18 versus 13 as you get to the higher doses. All of this is suggesting that the low doses seem to be fine for basically protecting us from blood-borne infections. I think if you have bacterium for example in the bloodstream it probably doesn't take very much antibody for that to be opsonized and phagocytozed but once you start getting chronic disease we need to start using much higher doses. The question then is why is that? Dr. Stiehm showed some data for some of the things that we think are important such as pneumococcus and H. flu. but in some ways we don't know. There is a dispute over whether mycoplasma is important. Moraxella catarrhallis was a commensal organism when I was a resident and now seems to be a significant pathogen. Are these other organisms responsible for causing some of these chronic infections, or is it simply a matter that it takes much higher levels of the other relevant antibodies, such as pneumococcus and H. flu., to just push it into these tissues where it is less available? For the most part we have relied on the data for IGIV products for the information that we are providing. To show you one representative table, every product is tested against a wide variety of antigens but the assays are sometimes very difficult to relate to what we do and look at in patients. Again, I am still not convinced that taking an IgG preparation that now has either a glycine or a sugar stabilizer and diluting that out gives you exactly the same titers as the assays that have been standardized in plasma or serum for patients. This is a very busy slide and I am sure you can't see it but you have a copy in your handout. But I just wanted to show that, you know, at least we think we have good titers for things like H. flu. and pneumococcal serotypes. There is some variation from brand to brand and there is also variation from lot to lot but in general we think these titers are very good. But for pneumococcus, I just wonder in some ways. If you go back to Jerry Schiffman's original data--and these are the pre-immunization titers for healthy adults and his assay is a little different so what he did is he looked at this, and he averages about 300 ng/mL in a normally healthy person. In fact, the so-called protective level is really based on the fact that healthy people have levels like that. It was never really based on what we can do in animals, and that is give a certain amount of antibody and then challenge them. So, we have never really done challenge studies in people and I doubt that the FDA would really approve of that. But I just wanted to make the point that most of what we know about protectivetiters is really done by inference as opposed to direct study. Virtually every study that we have done has really focused on IgG levels, and it is a good surrogate marker. It does tell us that we are replacing IgG but we have really not looked very much at specific antibody. There have been a few trials. We did a trial where we focused on looking at pneumococcal and tetanus half-lives, and we published the half-lives but we really didn't focus very much on how well we were protecting the levels. In fact, I want to go back and try to find that data and see if maybe we can really look at it in terms of what the absolute antibody titers were as opposed to what the half-lives were. There are very few studies that have actually published titers. They may have done the half-life studies but they haven't published the titers. There are a couple of studies that have used opsonophagocytosis assays that do suggest protective levels in patients but, again, it is a little hard to relate that back and we have never done good studies where we have actually followed the titers. I think that Dr. Stiehm was suggesting that perhaps we need to do that in future studies and really focus in more on protective antibodies. This is one study that did look at antibody titers by either neutralization or CMV but these patients were on very high doses, 500 mg every 2 weeks. It is also 1992 and, you know, there is a suggestion that the titers at least titers may not be representative of what we can achieve. So, in conclusion, I think the clinical data strongly suggests that higher doses of IGIV are more effective. I am not sure that we have even shown that we have max'd out on this. It does get to be a practical issue. Getting back to the question of how often you give it, if you do this more often there are a few things that happen. One is your peak and trough would rest to the mean. The other things is that we know when you give very high doses and when the level is high the catabolism is much higher; it is dependent on dose. So, if you give a very high dose your catabolic rate is higher and it is actually less efficient. So, if we did that same dose that they did in The Netherlands and gave this every week, the same dose but divided every week, the trough levels probably would have been significantly higher and maybe we could get them at the same dose up to 1200 mg. But for a lot of these we need to know what level of antibody is protective, and not only what is protective to keep us from getting an infection but what do you do once you are infected because, indeed, we are not only preventing infections in these patients but often having to treat them. At least for pneumococcus, I have seen the specific antibody disappear within 2 days when a patient became infected. Again, what are the most relevant organisms? We focused in a lot on the ones that patients seem to get with acute infections but what about things like Moraxella catarrhallis or mycoplasma? Are these important? Again getting back to what Dr. Stiehm said, are there other ways to determine this besides doing the big clinical studies? The largest study ever done still had difficulty showing differences between the two products because it didn't meet statistical significance in any area and in the general infections it didn't. I don't think anybody is going to do any study larger than that. So, we do need other ways to do this to decide what antibodies we are going to look at. I think after the break we are going to have a panel discussion and see if we can address some of these issues. Thank you. DR. SCOTT: Let's take a 15-minute break. [Brief recess] Panel Discussion DR. SCHIFF: Welcome back, everybody. In the next 20 minutes we are going to have a panel discussion to go over some of the issues that were raised in this morning's talks. Twenty minutes isn't nearly long enough. I remember when I was taking a biochemical chemistry exam and my professor came by and just kind of glanced at my paper and he said, "well, it looks to me like you know what you're talking about today." I said, "I don't understand; you couldn't possibly have read my answer." He said, "no, your answers are short and your handwriting is neat so I know you know what you're talking about." It was a good lesson to me. I think the less we understand the more we talk about it. But this morning we are going to talk about it for only 20 minutes. Some of the things that I would like to try to address are the issue of the IgG level. I think we all agree that the IgG level is important but what trough level is important? Then, the other issue is, is the trough the only important thing? Is getting a high peak to really force the IgG antibodies into the tissues important? Is that of any relevance? Or, is just maintaining a high trough important? We certainly should address the issue of antibody titers; which titers we are going to look at; which things are relevant for the patients. Dr. Stiehm outlined some of them. I brought up the issue of mycoplasma and Dr. Gelfand said he has looked in his IgG preparations and there is no mycoplasma antibody. So, it doesn't sound like we can raise the dose high enough. Zero times anything is still zero. So, there are issues. IgG doesn't cure everything and we need to sort of decide how we can focus and determine how we can do studies that are practical to answer some of these questions and, ultimately, I think also who is going to support them. So, why don't we start off with this issue of the IgG level, what levels are improvement or peaks are important? Hans, do you want to start with that? DR. OCHS: One problem that I see is what is a trough level really. What is important is what is above the baseline. So, the trough level alone from one patient to the next is probably not the absolute criteria because you may have a patient with X-linked agammaglobulinemia who has a baseline of 50 mg/dL and you may have a CVID patient who has a baseline of 450 mg/dL or 500 mg/dL. So, you would like to have one at least 500-600 mg/dL for the XLA patient. For the CVID patient you want to have above 1000. So, my trend is to look at the baseline which is essentially the IgG level prior to treatment with IGIV. That is not so simple because we don't know if over the years that baseline changes either up or down. So, the ideal trough level is something that we have to make an effort to form an educated guess for, but that is one thing we have to consider. The other is, you know, does the trough level reflect what the extracellular level is? For most patients that is important, in addition to what the surface level is on mucosal membranes. But, certainly, the peak level after the injection, the infusion of IGIV, represents only what is in the blood but not what is in the tissue, and the tissue level is also important. I think the tissue level is best estimated by the trough level. By that time there should be equilibrium. That brings us, of course, to the question of should we maintain a high trough level like we can achieve by giving either more or more often, every day or once a week? Subcutaneous gives you a much higher trough level than IGIV. So, these are all issues which we need to address and it depends on our clinical perspective. DR. SCHIFF: Does anybody on the panel or anybody in the audience have data on antibody levels in tissue after infusions? Has that ever been addressed, especially antibody titers in tissue? PARTICIPANT: I think it is very difficult to measure in tissues. By doing careful pharmacokinetic studies you can get some idea of tissue distribution but I think one relatively simple way to measure that antibody is getting to the right place is to take mucosal samples. That is my bias is, again, from the HIV work that I am involved with but isn't it very important here too? And, isn't it a lot easier to measure and assess looking at mucosal samples in vagina, rectum, saliva, bronchial secretions, rather than trying to find out what is in the tissues? We haven't tried to get that data, and maybe that is something we should think about seriously. DR. OCHS: Way back when I was a Fellow I tested the effect of bacteriophage on the antibody levels also in normal controls and, doing that, I had hoped that perhaps we would get mucosal antibodies. I personally, and some of my co-workers, sniffed the stuff. We never got any local antibody production. But if we looked at saliva or tears after intravenous injection we could estimate that about a tenth of a percent of the antibody titer in the plasma comes out in the tears or in nasal washing. So, there is a tremendous difference between what is in the serum, as you suggested, and what is coming out in the secretions, at least in the eyes and the nose. From that point of view--you know, this is a crazy idea but maybe we have to give gamma globulin IgG locally either into the eyes as drops or as an inhaler. There is a very