Learn about all myeloma happenings on the new Myeloma Crowd site: the first comprehensive site for myeloma patients and caregivers. Dr. Stephen Harding The Binding Site Group Interview Date: July 18, 2014

 No myeloma patient wants to hear the word "relapse," but how can we identify it early so it can be treated? A new FDA approved test called Hevylite® is now available that can pick up myeloma growth before it is seen in the standard SPEP and IFE tests. This new test can also help identify minimal residual disease, even if patients are considered to be have a complete response (CR) or stringent complete response (sCR) using the traditional tests. A test called Freelite* was the first to help myeloma patients identify "normal" and "abnormal" free light chains (kappa and lambda) and the ratio between the two. The Hevylite test expands our knowledge to also tell us how the immune system is responding - is it still suppressed or is it bouncing back?  The Hevylite test looks at the heavy part of the chain (IgG, IgA, IgM), immunoglobulins which protect agains bacterial and viral infections. Even small amounts of abnormal light chains can be bad for the immune system and this new test can help patients tell how their myeloma is responding to treatment and how their immune system is recovering (or not). In this show, Dr. Stephen Harding of The Binding Site Group in the UK gives great detail about the use and benefits of the Hevylite test and how it is entering the clinic. This test is now available at the Mayo Clinic and LabCorp and its use is expected to become a regular tool in monitoring myeloma and in testing for minimal residual disease.  * For more on the Freelite test, the IMF has a very helpful brochure here. Watch for a new brochure explaining the Hevylite test on www.myeloma.org coming soon. The live mPatient Myeloma Radio podcast with Dr. Stephen Harding

Jenny: Welcome to today's episode of mPatient Myeloma Radio, a show that connects patients with myeloma researchers. I'm your host, Jenny Ahlstrom. Now, if you'd like to receive a weekly email about past and upcoming interviews, you can subscribe to our mPatient Minute newsletter on the homepage or follow us there on Facebook or Twitter and please share these interviews with your myeloma friends. We have a new site called myelomacrowd.org. That's the first all-inclusive site for myeloma. We invite you to contribute to this site. If you'd like to contribute a post on your experience or your perspective with myeloma, click on "Become a Contributor". We've just added all the Mayo Clinic and Memorial Sloan-Kettering doctors to the myeloma specialist directory, so take a look at that growing directory. Today, we are very fortunate to have with us Dr. Stephen Harding of the Binding Site Group in the UK. Now, most myeloma patients know they have either kappa or lambda myeloma. This is detected with the Freelite test from the Binding Site Group, but testing the heavy part of the chain also may give us more information and may be very important in detecting minimal residual disease. Several patients requested this interview because we focus on the importance of diagnostics to get more personalized care. Dr. Harding, welcome and thank you very much for joining us on the show.

Dr. Harding: Thank you very much, Jenny. It's a real pleasure to be here.

Jenny: Well, let me introduce you for a bit. Dr. Stephen Harding is the Research Director of The Binding Site Group. Dr. Harding has spent the majority of his professional career developing monoclonal and polyclonal antibodies. These antibodies have been used in immunotherapies and immunodiagnostics. Dr. Harding is passionate about proteins and assay development and has been instrumental in the launch of Hevylite, this new IVD assay for the quantification of Ig kappa and Ig lambda serum immunoglobulins. In addition, Dr. Harding has been instrumental in launching approximately 20 other assays. Currently, he runs a team of 60 R&D scientists and a regulatory affairs department with a keen interest in 510(k) FDA submissions. Dr. Harding has contributed to many articles in peer reviewed journals and tens of published abstracts. Thank you again for joining us. Maybe we should start with just a little bit of background and history on your current role with the Binding Site.

Dr. Harding: Sure. I joined the Binding Site in 2006. My main responsibility was to organize the early clinical trials looking at the efficacy and the utility of Freelite. After around about six months, I was then given the opportunity to start developing an entirely new assay, the Hevylite assay, and I developed that from 2007 until launch, and the assay has just received FDA approval for IgG Hevylite, for IgA Hevylite, and IgM Hevylite. In 2011, I became the Director of Research and Development, and that role gives me the opportunity to liaise with myeloma physicians and myeloma experts and meet myeloma patients worldwide. I'm delighted to be able to have the opportunity to share with you our thoughts on the Freelite assay and the Hevylite assay.

Jenny: Can you tell us about the Freelite assay when that was developed?

Dr. Harding: Sure. The Freelite assay was developed and launched back in 2001. I think you were saying all myeloma disease is a kappa or lambda. I think if we go a little bit further back, myeloma is counter-associated with plasma cells and plasma cells produce antibodies. And we, as mammals, produce a huge array of antibodies. In the Anderson and Anderson publication in 2002, they said there were over a million different individual antibodies in the human body. These antibodies have a heavy chain. The most common is IgG. It could be an IgA or an IgM, and a light chain, either kappa or lambda. The light chain miring to the heavy chain to make a functional antibody is quite an inefficient process and the light chains are produced in excess, we think about 40% excess compared to the heavy chain. These light chains were able to be detected in the serum by the Freelite assay for the first time. The assay identifies a hidden epitope (the part of an antigen molecule to which an antibody attaches itself), so when you have your light chain and heavy chain paired together, there's an epitope that's hidden and it's only visible when the light chain is free. Over 90% of myeloma patients have an abnormally elevated free light chain and these are unique to the patient. As Anderson and Anderson showed us, there are a million different possibilities with your immunoglobulins, and so there are millions of different possibilities of your light chains, and these light chains are very individual. The Freelite assay uses a polyclonal antibody technology which is the only method that allows recognition of all of these unique molecules.

Jenny: That makes sense, so that's testing for a wide range of all the antibodies that your body produces, not just one. We'll talk a little bit about the monoclonal versus polyclonal I think a little bit later. Can you give us a little bit of history before we get started into the details about the mission and goals of the Binding Site?

Dr. Harding: The Binding Site is committed to improving patient lives worldwide and the way we do that is through a process of education, collaboration, and innovation. It's been my real pleasure to have worked with organizations in the US such as Mayo Clinic, Arkansas, Dana-Farber, Sloan-Kettering, the NIH, and support of clinical studies which look into the utility of our assays. I think as a sound bite, we may think about it in these sorts of terms. Individuals may beat cancer, which is absolutely amazing, but we believe together, collaborations between scientists, clinicians, laboratorians, and patients will actually conquer it and that's what gets me out of bed at 5:30 each morning.

Jenny: Thank you. Can you also share a little bit about your personal background? We talked a little bit before the show started, but I think others would be interested as well.

Dr. Harding: Sure. I'm a biochemist by trade. If during the course of our interview I start talking in too technical terms, please feel free to interrupt. I started my career making antibodies to the heat shock proteins, which are stress proteins and known to be elevated in several different diseases. Indeed, a treatment targeting HSP 90 has been evaluated in myeloma for some time. I moved to the University of Leicester to study a post doc looking at signaling pathways and again, I was making antibodies against p38 and JNK kinase signaling proteins. As I moved into biotechnology, I started making antibodies that were specific for different carbohydrate moieties on colorectal cancers. These antibodies were used in therapeutics and the company that I was working for at that time sold the antibodies to a larger company and I had the opportunity to join Binding Site. My career has really been spent in and around the association of proteins and antibodies, and how they can be used either as therapeutics or as diagnostics.

Jenny: Well, I would think with the really recent push in antibodies that you would be very excited right now.

Dr. Harding: Absolutely. I think in the setting of myeloma, the daratumumab and elotuzumab, really the early work that has been done with these monoclonals, it's very exciting. I've been fortunate to be able to attend meetings with clinicians that discuss these advances. The monoclonal antibody therapeutics or the idea behind monoclonal antibody therapeutics is not new. Back in the 1980s, we spoke about a magic bullet approach and antibodies have radioactivity or the chemotherapies bound to them. Really we were quite naïve at that time because the hypothesis and the delivery were somewhat disjointed. We are now much better at identifying the monoclonal antibody targets and this can be seen with the anti-CD38 target with daratumumab, of course. And so, these are starting to have genuine therapeutic uses. I think the most famous antibody, of course, is rituximab, which has revolutionized CLL patient treatments. My real hope is that the current batch of antibodies that is being evaluated in myeloma has a similar impact in patient outcome.

Jenny: Well, maybe your test will help determine that, so let's talk about your tests for a minute. We always push patients to ask about the test, the important tests that they should be getting. I think the Freelite test pretty much everyone gets, but I think there are patients that might still not know exactly what that test is telling them. Can you give us a lay person's description of first the Freelite test and why it's important and what it tells you as a patient?

Dr. Harding: Okay. The Freelite assay measures the amount of kappa light chain and lambda light chain in serum. As we're discussing earlier, these light chains form parts of your immunoglobulin molecule which are produced in excess. The light chain assay was revolutionary because for the first time, it allowed us to quantify what a normal kappa and lambda light chain component look like in a healthy human serum. Looking at that -- and this was worked with the Mayo Clinic -- we were able to develop a normal range for kappa and for lambda on a normal ratio of kappa to lambda, which allowed us to then evaluate where there was an abnormal production. Of course, before you can know what abnormal is, we have to have a very well described normal population that was completed by Prof. Jerry Katzmann on 282 normal human sera. They then evaluated the test in disease states and the first disease that was looked up was nonsecretory myeloma. This was worked by Prof. Mark Drayson from the MRC and he showed that in a disease that had previously been thought to be producing no monoclonal proteins, the Freelite assay allowed the detection of monoclonal proteins because of the disturbance in the kappa to lambda ratio. The assay was then evaluated in AL amyloidosis and in fact, it was in AL amyloidosis that it received its first international guidelines and that was back in 2005 with a consensus opinion for AL amyloidosis being published by Morie Gertz. I think that the real sensitivity of the assay is in being able to distinguish disease from normal and the reason why this was not possible before is because previously, the monoclonal light chains had only been able to be detected in urine. In fact, it was the detection in urine many, many years ago by Dr. Bence Jones that led to the nomenclature and people still use this nomenclature as Bence Jones proteins.

Jenny: Now, as you have been in this space for a long time how did you begin to develop the Hevylite chain test and what was the process to go through that?

Dr. Harding: Hevylite, we were first looking at how Freelite has provided the utility in the clinical chemistry setting and the individual assay is either kappa or lambda that provided some utility, but the sensitivity of the assay was in looking at the ratio of kappa to lambda. It's the understanding that your kappa could be either monoclonal, so a single clone producing kappa, and the level of your lambda that gives you an indication of your polyclonal or normal immunoglobulin repertoire. And so, we thought that if we were able to provide a sensitive understanding of the patient's light chains using this kappa-lambda ratio, if we were able to advance stuff and start looking at an assay that is an IgG kappa and then after that, measure IgG lambda, then you would be able to have or produce rather an IgG kappa-IgG lambda ratio, and that may be more sensitive than the current technologies. We would then move this to IgA kappa and IgA lambda, IgM kappa and IgM lambda. The assay used similar to the Freelite assay uses polyclonal antibodies because the immunoglobulins that we're recognizing are all individual and we have targeted an area in the immunoglobulin in what we'd call the constant region of the immunoglobulin. And so, the assay recognizes all possible immunoglobulins, which means we can monitor and measure all possible myeloma or monoclonal immunoglobulins. It uses a pool of myeloma proteins as an immunogen and a larger pool of myeloma proteins to purify, and each of our batches of Hevylite are thousands of immunizations and thousands of different proteins in size.

Jenny: Well, let me ask a question. When you talk about the ratio between kappa and lambda, I've had some doctors say you should really not necessarily consider the different levels of the kappa or lambda when you're looking at the Freelite test, but just focus on the ratio. And then I've had other doctors say no, you need to look at the ratio and the other level, the actual level of the kappa or the lambda. I'm not very clear on which is the right answer, so maybe I should ask you because you are the creator of the test.

Dr. Harding: This, in fact, is a very common question that we get asked. The kappa-lambda ratio is a very useful marker of clonality, so it is able to distinguish between normal and monoclonal. In the assessing of a screening tool, one would use an abnormal kappa-lambda ratio to identify a patient with perhaps monoclonal process. In monitoring the patient, you would look to use either the involved, i.e. the clonally produced kappa or lambda, or a difference between the involved and the uninvolved, so you would use the absolute measurements. Somewhat confusingly, as the patient has a very, very good response and has a complete response, we then go back to looking at the kappa-lambda ratio in order to assign what is called the stringent complete response. So I'm afraid the answer is we use the ratio at presentation and at maximum response, and to monitor the patient, we use the monoclonally produced light chain.

Jenny: Well, that makes sense, so the answer is they're both right.

Dr. Harding: Absolutely. The sensitivity of just using the kappa or just using the lambda without the ratio would be limited. It would still be a very useful test, but it would be much more limited. The ratio is really an exquisite tool of identifying monoclonal processes at presentation. Indeed, the work by Jerry Katzmann in Mayo Clinic suggests the algorithm of Freelite looking at an abnormal ratio on certain protein electrophoresis is the most cost effective algorithm for identifying all monoclonal gammopathies.

Jenny: So the Freelite, when you talk about minimal residual disease -- and I know we'll go on to talk about that later in the show, but just to address it a little bit, the Freelite test would be pretty instrumental then in monitoring minimal residual disease if the ratio you're talking about is how well you've responded basically. Is that correct?

Dr. Harding: Absolutely. The International Myeloma Working Group have identified that the free light chain ratio alongside flow cytometry at maximum response can be used to assign stringent complete response. This is a greater depth of response than just looking at immunofixation negativity, which was the standard assessment of complete response. Earlier this year, Kapoor from the Mayo Clinic published that the achievement of a stringent complete response was associated with a superior outcome compared to standard immunofixation response. The International Myeloma Foundation have got a cure myeloma initiative, the Black Swan Research Initiative, and they have included both Freelite and Hevylite as markers of response alongside flow cytometry path, et cetera, and has markers of depth of response. There have been several publications at ASH to the European Hematology Association meetings which have shown that the depth of response measured by Freelite and Hevylite may be superior to standard serum electrophoresis responses.

Jenny: Well, that's good to know, so let's talk a little bit more about the new test. I'm not sure that I still understand exactly how it works or what it tests for and what it tells us, so can you go into a little more depth about that test?

Dr. Harding: Each individual components of the Hevylite assay identifies an intact immunoglobulin, heavy chain and its light chain partner. So in myeloma terms, you may have heard patients say that they have an IgG kappa myeloma and this is because they have an IgG heavy chain and a kappa light chain. The assay quantifies the IgG kappa and a second assay would quantify the IgG lambda. And so, you are able to get a specific IgG kappa-IgG lambda ratio. Why this may be important is we were talking earlier about the light chain ratio being a very sensitive marker of disease at diagnosis and a very sensitive marker at complete response because it gives you an indication of both the production of the monoclonal immunoglobulin, the disease state of immunoglobulin, and also the normal polyclonal immunoglobulin, and the Hevylite chain assay builds on that relationship. Where it may be used is that monitoring patients with myeloma can be very difficult for laboratorians and this is because current assays such as serum electrophoresis can be interfered with by other proteins. The IgA monoclonal protein migrates into a position where there are numerous other serum proteins and this can make life very difficult for the laboratorian. We call it co-migration and it can occur in between 30% to 50% of all IgA monoclonal proteins. In addition, the monoclonal proteins may be very diffused and difficult to identify. And so, we know that the changes in monoclonal protein load give the physician a really great insight into whether the patient is responding to treatments or not. If we are not accurate in our quantification of those changes, then we may either have an underestimation or an overestimation of response. The Hevylite assay does not have the same interference. And so, it gives a very accurate measure of the monoclonal protein load.

Jenny: So it's determining response levels again at a deeper level. If the ratio is important -- and I know there are a variety of combinations, so you can have IgG kappa or lambda, or you can have IgA kappa or lambda, or you can have IgM. I don't know how many that totals, but there are like eight to ten combinations that you can have as a myeloma patient. Does that test give you any feel for the better or worse prognosis? Most people have said to me being IgG kappa or IgG lambda or IgM or IgA doesn't necessarily give you prognostic information like del(17p) or something like that would. Can you weigh in on that at all? Does it or is that accurate?

Dr. Harding: Absolutely. Having an isotype which is either an IgG kappa or an IgA lambda is not associated with an adverse outcome, so the intact immunoglobulin in isotype is not associated with an adverse outcome. Having an abnormal Freelite is associated with an adverse outcome and this was worked from the MRC. What we have found is that using the Hevylite assay, if you look at an IgG kappa patient and they have a very extreme IgG kappa-IgG lambda ratio, or if you look at an IgA lambda patient and they have a very extreme IgA kappa-IgA lambda ratio, then these patients have a poor outcome compared to those patients whose ratio is not so extreme. And so, while the absolute isotype doesn't predict outcome, the relationship between the tumor-produced immunoglobulin and the polyclonal normally produced immunuglobulins is actually highly predictive. There were two papers, one by Prof. Jo Bradwell and the other by Prof. Heinz Ludwig who suggested that the Hevylite ratio and actually the Freelite ratio as well can give you an indication of the severity of disease because it gives you an indication not only of the amounts of tumor because of the amounts of immunoglobulin being produced, but the impact to that tumor because of how much suppression there is associated with the presence of that tumor. So del(17p) and t(4;14), absolutely adverse risk markers, isotype, IgG kappa, IgA kappa, IgG lambda isn't associated or doesn't prognosticate in these patients, but understanding the relationship between the monoclonal protein and its polyclonal counterpart, that’s highly informative.

Jenny: Okay, so if I had to restate what you just said in lay patient terms, I could say that these tests are trying to determine how much tumor is being produced and what it's crowding out because of how much tumor it's producing. Is that correct? Is that how you would state it?

Dr. Harding: Absolutely.

Jenny: Okay, and you can add to that.

Dr. Harding: Sure. It's not just in how much tumor is or whether the tumor is crowding out rather. Some tumors might produce quite small amounts of monoclonal protein, but have a very big effect on the polyclonal, the normal immune system. And so, we don't understand the relationship between the tumor and its impact on immunoparesis. Actually, if we were to work backwards into monoclonal gammopathy of undetermined significance, actually in MGUS patients, comparatively few have immunoparesis (lowered blood immunoglobulins), but those patients that do have immunoparesis and those patients that have immunoparesis measured by the Hevylite assay have an increased tendency to transform into myeloma, so the tumor can have an impact irrespective of its bulk, and that's something that's very new and exciting about the Hevylite assay because it's giving us a hidden insight into the biology of monoclonal clones.

Jenny: That makes more sense. So when you talk about minimal residual disease, and we've heard that term a lot, but I think a lot of patients may not completely understand the importance of it or why it matters, so can you give us your take on why minimal residual disease is so important?

Dr. Harding: If we were to go back into the history books, a complete response in myeloma was a greater than 70% reduction of the monoclonal protein. And then as we have improved tests in the immunofixation and electrophoresis test, then we can start to say that a response is a clearanceof the monoclonal protein by immunofixation. An increased sensitivity and the ability to detect for the tumor would then be the Freelite assay and then we say actually the better response than just having a negative immunofixation is by having a normal free light chain ratio. We can add to this by then looking at flow cytometry and say if we can't detect any tumor cells by flow cytometry, this is a step further in our understanding of how much tumor is left or whether we have eradicated the disease. You see, minimal residual disease and response is relative to the test that you are using. If your test is insensitive, then the amount of tumor that you cleared is indicated only by the sensitivity of that test. Excitingly, we are in a phase with our technology where we have very, very, very sensitive tests to identify very small numbers of monoclonal clones and this can either be done with the serum tests and the Freelite assay, and the Hevylite assay, the flow cytometry, or by looking at imaging. It seems a very obvious statement, but the less tumor you can identify by the most sensitive technique, the better the patient's outcome will be.

Jenny: I don't think I really completely understood it until I had a conversation with Dr. Landgren who was talking about minimal residual disease. When he explained that it matters and it's relevant because it depends what you do next -- for example, he was saying you could take a patient who had an extremely good response in all these tests, the Freelite, the Hevylite, the flow cytometry, the imaging and then you're measuring response with earlier tests and then you know what you do next, how you treat the patient. Do you go for a more aggressive therapy because they're not responding well or less aggressive therapy because they are responding well? Once he started talking about it in those terms, the light started going on for me about why minimal residual disease is so important because then they can craft a very specific plan and it seems that many myeloma patients have been treated in much the same way over the last 20 years, I would say, with the same drugs and the same approach. It just seemed like it opened up a whole new window to a new method of treatment.

Dr. Harding: Absolutely. I was speaking with Dr. Landgren after his ASH presentation last year. He presented the changes in the monoclonal protein using the Hevylite assay, were the most sensitive serum markers or serum indicators of response compared to standard electrophoretic techniques. When speaking with Dr. Landgren at ASH, he was saying that for the first time, myeloma doctors have a choice because of the next generation of treatment, the proteasome inhibitors in Velcade, the monoclonal antibodies, Revlimid, etc. There's now a whole plethora of different treatments that are targeting different aspects of the myeloma and its surrounding environment, which means that the tests that you use to identify whether a patient is responding earlier or whether a patient is not responding give you an opportunity to then change that treatment, so I agree wholeheartedly with Dr. Landgren.

Jenny: Well, I would have to add to that and just say they can only change the treatment that you have if you are working with an expert myeloma doctor because there are many doctors that may not know what to do with that additional information even though it gets some people like you and some of the myeloma specialists very excited to have that level of detail. Unless you have a doctor that knows what to do with that information, they may or may not be able to act on it.

Dr. Harding: I think one of the commitments from the International Myeloma Working Group is to provide guidance to all doctors on how to use tests and how to assess minimal residual disease and the opportunities around therapy. They provide consensus opinions and publish these on a routine basis.

Jenny: They just had their latest meeting in Milan, Italy, which was very helpful. So about this test, now that it was FDA-approved, where is it available? How would a patient access it? Is it being used regularly in the clinic? How does a patient know if they've had this test or how to ask for this test?

Dr. Harding: I believe that in the USA, Mayo Clinic are using the test and I think LabCorp are about to start to use the test. There are numerous sites that have been evaluating the tests including the NIH and the sites we mentioned earlier. Because the assays were only recently FDA-approved, it's not available in all facilities, but I think part of the improvements or part of the improvements in patient's understanding is that patients and physicians should be able to discuss the merits of how their protein is assessed and then identify the laboratories to run the test. In Europe, the test is used in Germany and in the UK and in sites in France and Italy.  Well, I think it's about whether the laboratory and the physician have the discussion on how they want to monitor the patients, whether you would know the Hevylite or the Freelite assay had been used.

Jenny: If you're measuring the presence of a tumor burden or the amount of tumor burden, I would think you would need to use the test multiple times at different stages in your therapy. Is that true?

Dr. Harding: Every clinical study and physician has their own protocol. Of course, patients and physicians should discuss how often they should be monitored during the course of their treatment. Essentially, every time a patient is monitored, the Hevylite or the Freelite assay could be used. In the same way, one might order electrophoresis or immunofixation, but it's very dependent on whether there is a clinical trial protocol or how the patients and the physicians are discussing their approach to their monitoring.

Jenny: And just to put it in perspective because you've already given us the purpose of it, but if you give it in perspective with the other tests like the gene expression profiling, these might be done once. If you are monitoring -- let's say you've gone through your course of therapy, you'd want to monitor it beforehand and after, so it just seems to me just from listening to you that it might be something you would want to ask your doctor about regularly. Now, is this an expensive test to request and do you know if insurance pays for this test?

Dr. Harding: Currently, I don't think that we have reimbursements. I am not entirely sure. The cost of the tests, I'm afraid -- I am Director of R&D, so I don't actually know in different countries how much the tests would cost, but this is a comparatively cheap test compared to, for instance, gene profiling, which is thousands of dollars. This is certainly not in thousands of dollars. It's in the over $10 test, for instance, but a very loose figure. With respect to frequency, myeloma patient's disease is monitored by blood tests on a routine basis. The way patient's responses are assigned is by changes in these blood tests. This is the International Myeloma Working Group's response assignment. If one had a very good partial response, that would be a greater than 90% reduction in the monoclonal protein as measured by serum protein electrophoresis from presentation for the points of evaluation. The Hevylite test can be used in exactly the same way. There was recently a publication at the IFCC that showed that changes in the Hevylite value could be used in exactly the same way as changes in the monoclonal protein measured by serum protein electrophoresis. It's a very easy-to-use test and it doesn't require bone marrow aspirates or prolonged imaging. It's a simple blood draw. It's actually quite an accessible test, too, in the same way that the Freelite assay is a very accessible test.

Jenny: You mentioned, you began this test just testing the IgG, which is the most common type of heavy chain, but now that its available for IgA and IgM, or is it just IgG for now?

Dr. Harding: We have FDA approval for IgG, IgA, and IgM. The FDA approval for IgG and IgA is -- actually, it's a test that's approved to monitor myeloma patients, so to monitor the changes in monoclonal protein in myeloma patients. The IgM test is approved only to measure the protein. That is because we have at the Binding Site done substantial clinical studies to compare the utility of the IgG and IgA assays in monitoring myeloma patients compared to standard tests, and we are very pleased that the FDA agreed that this assay provided a sensitive tool for monitoring.

Jenny: Now, with your expertise on polyclonal antibodies -- that's your background -- and monoclonal antibodies, can you give us your opinion about these newer options? You've mentioned it at the beginning like daratumumab and elotuzumab in myeloma. Can you tell us where you see that future heading and have there ever been any polyclonal antibodies developed which are targeting multiple antibodies? I know they're starting with monoclonal, but do you see it ever being expanded?

Dr. Harding: Perhaps it might be worthwhile explaining to the listeners what the difference between a polyclonal and monoclonal antibody is, and then to explain why therapeutically we choose monoclonal antibodies, but in an in vitro diagnostics, we generally choose polyclonal antibodies.

Jenny: That'd be great.

Dr. Harding: Polyclonal antibodies are mixtures of antibodies that recognize different epitopes. We started today by discussing plasma cells, and plasma cells produce antibodies. And we, as human beings, we produce thousands, tens of thousands and hundreds of thousands of different antibodies against bacteria, and each of these antibodies recognize a different epitope. That's called "poly" which means many; "clonal", many different clones, many different antibodies recognizing many different epitopes. A monoclonal antibody is where we isolate a single clone and we remove it from the rest of the clones and we immortalize it in a hybridization process, and then that plasma cell becomes an immortal plasma cell, but it only ever produces a single antibody. Now, historically polyclonal antibodies were used in diagnostic tests because in the diagnostic test, you want your diagnostic test to recognize as many different patients as possible. If we think about the Freelite assay for a moment, Anderson and Anderson said there are million or more different immunoglobulins in the body, which means there are a million or more different light chains present within the body. That means that you want a test that recognizes as many of those different light chains as possible because each of them will be unique and you need a lot of recognition to make sure you cover all of the possible epitopes and all of the possible amino acid sequences within those light chains. If you were to use a monoclonal antibody, you would only recognize a single epitope or a single sequence within the light chain and that may or may not be present in the light chain that you are testing. So from an in vitro diagnostics point of view, you use a polyclonal approach, which is to identify as many as possible. Therapeutically, you actually want to use the monoclonal approach and that is because you want to use an approach where you have the very strong control over the epitope that your antibody is binding to -- for instance, anti-CD38 or antic-CD20 -- and you want to be able to reproducibly produce this antibody because you have to grow it as a clone and you perhaps can do that from a polyclonal approach. And so, if we think about whether polyclonal antibodies may ever be used in therapeutics, actually, polyclonal antibodies are used in therapeutics if patients are given IVIG, are given intravenous immunoglobulins, that is polyclonal antibodies that have been taken from healthy blood donors, and monoclonal antibodies are where we use a specific antibody to target a specific epitope to give a specific response, so we do use polyclonal antibodies. They're not quite used in quite the same way as the anti-tumor antibodies, but certainly are useful therapeutic options.

Jenny: It sounds like you're testing for everything and the monoclonal antibodies like daratumumab or SAR or elotuzumab that are out right now are targeting one specific thing like either CS1 or CD38 and those types of things.

Dr. Harding: Absolutely. In an in vitro diagnostic terms, it's very important to use a polyclonal approach because your antibodies in your light chains will be as unique as we are. From a therapeutic point of view, you want to identify a single clone or a single target to be able to reproducibly use it as a drug, so the two systems are very different.

Jenny: Well, thank you very much. Now, I want to open it up for caller questions because we have some callers on the phone. So if you have a question for Dr. Harding, please call 347-637-2631 and press 1 on your keypad. Our first caller question, please go ahead.

Caller: Hi, Dr. Harding! Thank you for taking the call. Question, I listened to the show and as patients and caregivers, this is still pretty complicated stuff for us. I understand the importance of a stringent complete outcome. Now, the previous tests can identify stringent or complete outcome. Can you help me understand how this test that you're talking about, this new Hevylite test, is an additional indicator for this stringent complete outcome? Is there really additional information that you're gathering and what's the benefit to us as patients?

Dr. Harding: Thank you very much for the question. It's a really great question. The standard assessments of complete response using immunofixation has a sensitivity of about between 200 mg/L to 500 mg/L. It will only tell you whether your monoclonal component has fallen below that level. The Hevylite assay will give you an indication both of how your monoclonal protein has reduced, but will also tell you about the recovery of your polyclonal proteins. The recovery of the polyclonal gives you an indication that a healthy immune system has started to recolonize or to regrow after treatments.

Caller: Okay, so the difference is in my previous tests just tell me if there's no cancer. This test tells me if I have a healthy regrowing immune system, so the recovery of my body after the disease.

Dr. Harding: Absolutely. The monoclonal components or the involved Hevylite will tell you if the cancer immunoglobulin has reduced on the polyclonal component, which is a new insight into myeloma biology. It seems to give an indication on whether the immune system has regrown and whether the patient has responded. Now, it's worth pointing out that there have been publications which support this hypothesis, but this has not yet been shown or been included in any clinical study. And so, it's very much an experimental observation.

Caller: But if a patient wanted to get as much information about a disease, this is helpful. Does this give me some kind of an indication of how my cancer is trending? What's the hope here? I know you can't say, but is the hope that it's showing me that I am not only in remission, but then my body is heading back towards a normal state? What am I hoping to figure out at the end of the day on this test?

Dr. Harding: The Black Swan Research Initiative has used Freelite or has included Freelite and Hevylite alongside standard assessments of response because we believe it's a whole battery of tests that will help us understand the complexities of myeloma, and myeloma is a hugely complex disease. I think what you're hoping to see is that in very simplistic terms, one would anticipate if you have a normal kappa-lambda ratio by Freelite and a normal kappa-lambda ratio, an IgG kappa-IgG lambda ratio, IgA kappa-IgA lambda ratio, et cetera, then that gives you an indication that your body has returned to, as you say, a more normal state. We're still involved in the experiments of clinical studies to prove those hypotheses, but that's certainly what we're looking for. We're looking to see whether the test adds value in giving an indication both of the reduction in the tumor load by the monoclonal protein reduction and also in the immunological recovery by looking at the polyclonal and immunoglobulin elevation.

Caller: Is this part of an ongoing medical trial that patients can participate in or is the trial already completed and approved by the FDA?

Dr. Harding: It's an in vitro diagnostic test, so we don't have the sort of trials that you're talking about in the same way that you have a drug trial where you can enroll, but there are a number of clinical studies in sites around the US and Europe which are evaluating the test and a number of sites that are evaluating the test including Mayo Clinic, NIH, Dana-Farber, et cetera. I think if you were to contact the Binding Site office in San Diego, they may be able to help or direct you to where somebody is able to have the test run over in the United States. The FDA have looked at the data that Binding Site submitted, which compared the use of the Hevylite assay to standard methods of monitoring patients and their approval is that they found that the assay had been proved to be substantially equivalent to the standard method, which is why we received our 510(k) approval.

Caller: Okay. My last question is as a patient, help me understand how do I talk to my doctor about getting this test. Will my doctor be okay with that?

Dr. Harding: As a biochemist and somebody not over in the United States, all  I would suggest is that you could talk to your doctor about the method that he's using to monitor the monoclonal protein or your monoclonal immunoglobulin and then there are publications that have been presented at ASH and in Leukemia Journal alongside educational materials from the Binding Site in San Diego, which could then be used to share and discuss with your physician.

Caller: Jenny, can you provide those links to us so we could use those with our doctors?

Jenny: Oh, sure! I'd be happy to. I'll include it as part of the transcripts.

Caller: Thank you. Thank you, Jenny and also thank you, Dr. Harding. I appreciate you taking the time with us and answering dumb questions.

Dr. Harding: These are very good questions. Thanks very much.

Jenny: Thank you for your question.

Caller: Hi, Jenny! Hi, Dr. Harding! It's a pleasure to speak to you. This is Dana Holmes and I have a question for you -- or two actually. I heard that you noted that those with immunoparesis have the increased tendency to progression. I'm smoldering multiple myeloma and I have some immunoparesis that I know of from the quantitative immunoglobulin tests that are done on me in my IgA. I'm IgG kappa and I'm presuming that my IgG is equally suppressed because that's where my monoclonal protein is found. So I would imagine this type of test would be very valuable for me to establish my risk to progression. I realized that some centers are currently using it in the US, as you noted, but I'm wondering if many of the clinicians are still looking at it at this point as more of an investigational tool and not yet ready for general use. Could that be a potential reason why we're not seeing it being moved into the clinic for every day use at this point, particularly for someone in my situation?

Dr. Harding: Thank you very much for your question and it's a pleasure to talk to you. I know we've exchanged some tweets.

Caller: Yes. Yes, we have, Dr. Harding. Yes. Thank you so much for that.

Dr. Harding: You're right. The assay is only recently launched in the US from FDA approval. And so, it's still very much in the experimental phase or the investigational phase, although I think that the work that's come out of Mayo Clinic and out of the NIH is showing good, genuine utility. There have been no studies looking at smoldering myeloma with Hevylite, although I'm sure you know that the Freelite assay is very important in smoldering myeloma as a marker of progression and certainly ratios above 100 are considered to be very informative. Unfortunately, I don't think there's enough evidence or enough work that's been done in smoldering to understand whether the assay has a utility in that specific indication or not. What we do know is that in assessing a monoclonal gammopathy, the level of suppression that was evident, it was a marker of transformation into myeloma and we do know in myeloma the level of suppression is an indication of the severity of the disease. What I think we should do -- and this is something that I'm certainly pleased to do from Binding Site -- is to see whether we can identify trials of smoldering myeloma patients or physicians who have a number of smoldering myeloma patients in order to start that experimental work.

Caller: I would welcome your support of that within the myeloma community because quite frankly, I think and I see it's a very important test. I would love as a smoldering patient to be able to access that whether it'd be done in a trial or otherwise, so I would welcome that and I'll keep my eyes open certainly for your tweets about it. I have another question concerning immunoparesis. If you have that at the start of treatment, can you eventually see immune reconstitution to normal levels after treatment or once the immune system is suppressed, that's it? It really will never bump back up and you'll see normal levels of the immunoglobulins, of the normal polyclonal immunoglobulins?

Dr. Harding: In looking at the Hevylite data that we've evaluated so far and in publications from Professor Heinz Ludwig from the Wilhelminen Hospital in Vienna, almost all or a large majority of myeloma patients present with some degree of immunosuppression using the Hevylite assay, but this immunosuppression recovers as the patient responds. And when the patient has a complete response or a stringent complete response, the levels of the uninvolved immunoglobulin seems to be approaching normal, so I think the immune system can recover once we have alleviated the burden of the myeloma tumor.

Caller: I see, and the Hevylite would be able to give you some insight as to how your immune system is actually recovering.

Dr. Harding: Well, that's what we hope. That's the indication of the data that's being presented to date. Binding Site has got a strong commitment into understanding and supporting patients with monoclonal gammopathy and trying to find out the best possible tests in order to evaluate the disease. We're very keen to understand what are the tests that may be useful in assessing immune recovery and that's an area of focus that I hope in the next few years, we may be able to be discussing other tests looking at immunoreconstitution.

Caller: Okay. Lastly, Dr. Harding, if you please, my question is related to the Hevylite assay and how it can be applied to a biclonal process, not so much the conventional biclonal, the IgA kappa to IgM lambda, but more on the sense of the same epitope. I personally have a biclonal IgG kappa. I'm showing two bands on IFE, but only one has a prominent M-spike, so really I only see the one M-spike, but I do have an abnormal free light chain as well. So I'm wondering, would the Hevylite shed some additional light on my disease process in any way?

Dr. Harding: Well, if I understood correctly, you have two IgG kappa clones?

Caller: Yes, and that's based upon the results from the immunofixation, the IFE. It clearly states that there is a second band.

Dr. Harding: Right, so the Hevylite assay would be able to identify both of those clones and would report it as a single value, i.e. the degree of IgG kappa that was present within the serum. Then the second test, the IgG lambda test, would be able to report the degree of polyclonal IgG lambda that was present in the serum. The assay can be used to quantify all of the G kappa and all of the G lambda. One of the areas where we're seeing some utility for Freelite and Hevylite to be run alongside one another is in assessing patients who have some clonal evolution. And for the first time, we're understanding that clones are a much evolving scenario rather than being a stable scenario. And so, I think evaluation of the G kappa, the G lambda, and the free kappa would give some indication of the disease process.

Caller: Okay, very, very good. Thank you so very much. It really was a pleasure. Thank you, Jenny, for again giving us this platform to speak directly to the experts. I appreciate it very much.

Jenny: Thank you, Dana, for your question. Okay. We also had an emailed in question by Susan. She says, "I have the kind of myeloma with no M-spike, non-secretory. Would this test be relevant for people with this kind of myeloma? Would it be better for follow-up than the usual kappa-lambda ratio test?"

Dr. Harding: That is a great question. We simply haven't looked at enough non-secretory myeloma patients to know whether it adds value. There's only been a very small study done in Leicester in the UK and there it seemed that 20% of the patients that had non-secretory myeloma had an abnormal Hevylite ratio, but we have no data to suggest whether it can be used to monitor these patients, so I'm afraid it's just too early in the life span of the test to understand its role. As we do know, of course, the Freelite assay identifies a number of these patients and is actually recommended in International Myeloma Working Group Guidelines as a tool to monitor non-secretory or oligo-secretory patients.

Jenny: Great, very helpful. Well, Dr. Harding, thank you so much for explaining this test to us. Sometimes this information can be pretty overwhelming because there are so many different ways of doing the diagnostics, so it's very, very helpful for you to explain how the tests work and what they do and why they're important, and I believe this is an important test, so thank you.

Dr. Harding: Well, thank you very much for the opportunity. The Binding Site has a commitment to continue education for physicians, patients, and laboratorians. And so, we produce documents for patients as well as doctors, and these should be able to be obtained from the office over in the United States. We appreciate how difficult this is and hopefully they provide some use for the patients.

Jenny: Well, it's very helpful and I will include it on the transcript, but we very much appreciate your research and the work that you're doing especially to create assays that are blood tests and not bone marrow biopsies. It's very helpful to us, so we are very, very grateful for your time today. Thank you so much.

Dr. Harding: Thank you very much.

Jenny: Thank you for listening to another episode of Innovation in Myeloma. Join us next week for our next mPatient ratio interview as we learn more about how we as patients can help drive to a cure for myeloma by joining clinical trials.