Fenghauang (Frank) Zhan, PhD
Yogesh Jethava, MD
University of Iowa
Interview Date: December 11, 2018

What cells cause myeloma and why do patients relapse after treatment? Myeloma stem cells may be part of the answer. 

Fenghauang (Frank) Zhan, PhD was recently selected as a Myeloma Crowd Research Initiative (MCRI) award recipient. The goal of the MCRI is to help optimize myeloma therapy for individual patients.

Dr. Zhan's noticed that patients whose myeloma is resistant to chemotherapy have higher levels of CD24 on their flow cytometry tests. He believes that the CD24 is present on the myeloma "stem cells" - cells that initiate the cancer and cause relapse. His work will validate that CD24 can indicate patients who are more prone to have resistant disease and is working with other researchers who are developing CD24-directed treatments. 

If patients were able to receive full myeloma therapy including therapy that could kill initiating cells, we would be much closer to a cure. Hear more in this show from Dr. Zhan and Dr. Jethava from the University of Iowa about this important research. 

The Myeloma Crowd created the MCRI because we “Can’t Wait for a Cure.” Through the MCRI we want to help find the right treatment for the right patient at the right time. The MCRI is funding three research projects – each with a different focus – that will also integrate with HealthTree, a new tool for myeloma patients. Learn more about HealthTree here.

 

Thanks to our episode sponsor

Jenny: Welcome to today's episode of Myeloma Crowd Radio, a show that connects patients with myeloma researchers. I'm your host, Jenny Ahlstrom. We'd like to thank our episode sponsors, Celgene Corporation for their support of Myeloma Crowd Radio. Today is the second show in a series of three on Myeloma Crowd Radio for the Myeloma Crowd Research Initiative or as we call it the MCRI. When we started the Crowd Care Foundation in the Myeloma Crowd, we asked our scientific advisory board what we could do to fill gaps in research funding. The first time that we did this, they told us we should fund high risk research. So we ran the very first ever crowdsourcing and crowdfunding campaign and we picked two immunotherapy projects.

The first was a CAR-T project from the University of Wurzburg in Germany that was targeting CAR-T cells with CS1 and BCMA. Another immunotherapy called MILs from Dr. Ivan Borello, an immunologist at Johns Hopkins. The second year we asked this question, Dr. Fonseca suggested that with all the new treatments, we should create a tool to help patients identity treatment options they could consider especially if they were being seen in the local oncology setting. Based on this need, we developed the HealthTree tool and with HealthTree patients can number one, understand their personalized of treatment of options to have better and more informed discussions with their doctors. Second, they can find clinical trials they're eligible to join based on their lab values and all the eligibility criteria. Third, they can share their myeloma experience anonymously with other patients and researchers to help identify faster cures.

In the summer, we canvassed 50 cities to show these tools with over 100 patients. Now, over 2200 patients are using HealthTree. And so we're very excited. We just begun showing some of the reporting in that with who is using HealthTree which is wonderful. Today, we'd like to announce a new feature in HealthTree that lets you write your personal myeloma story so you can share that later with your doctor or with other patients if you so choose. As we think about this, there is a lot of focus on precision medicine, but personalized medicine includes not only myeloma genetics and your lab and your personal experience with a disease like your family situation or the feelings about how you're dealing with this disease.

Now our focus for the second Myeloma Crowd Research Initiative is complimentary to what we built with HealthTree. We heard from doctors that a small fraction of patients could be cured, and we want to know who they are and how they're achieving success to become replicated for other patients. The second MCRI is focusing on optimizing myeloma care because so much can be done to link in life today with today's drugs if they're applied for the right patient and the right order at the right time. And we've asked all three projects to integrate in some way with HealthTree. Last week, we heard from David Chung from Memorial Sloan Kettering on his testing of immune system to create an immune system signature.

Today's show is about identifying personal strategies to prevent relapse or how to kill all the myeloma from the beginning. On Friday, we'll be sharing our third project, the 3D tumor modeling. Each of these three projects bring a personalized approach to caring for you as a myeloma patient. We'll be funding these projects for a total of $500,000 or more and invite you to join us. You can go to http://give.crowdcare.org/mcri and create an individual or team fundraising page then share it in any way you want. Join a local run or walk and share your page, share one of these interviews on social media during the holidays as a way people can support you in your efforts to just speed up a cure for myeloma. You can share it by email, Twitter, Facebook or other social media accounts. We know that your friends and family want to know how to support you during your disease journey and this is an excellent way of doing that. The end of the year is a great time to doing that as well.

Now, we would like to welcome two investigators to the program today, both at the University of Iowa who are working in tandem to make this research a reality, Dr. Frank Zhan and Dr. Yogesh Jethava. Welcome to the program.

Dr. Zhan: Hi. Good morning. This is Frank Zhan, I'm ready to get started to discuss my project with all of you.  

Dr. Jethava: Good morning, this is Yogesh Jethava. I am as well excited to discuss the project and the future prospects.

Jenny: Well, let me introduce you both before we get started. Dr. Frank Zhan is Professor of Medicine at the University of Iowa in the Hematology Bone Marrow Transplant department since 2012. He's also adjunct professor of the College of Dentistry in Dental Clinics since 2016. Previously, he performed research at the University of Utah for myeloma at the Huntsman Cancer Institute as well as the University of Arkansas for Medical Sciences, or UAMS. He helps review grants to identify those that should be approved for several foundations including the National Cancer Institute for R01 and R21 grants. These are the largest and most prestigious grants from the NCI.

He's also on the editorial board of journals like International Blood Research and Reviews, the American Journal of Blood Research, the Journal of Blood and Lymph and BioMed Research International. His current grants include the role of NEK2 in myeloma residual disease, genetic studies of myeloma genes, high risk myeloma approaches and the use of the vitamin C infusion in myeloma and what we'll be talking about today, the identification of myeloma stem cells. Dr. Zhan has very deep research expertise on myeloma genetics having over two decades of study in that field.

Dr. Yogesh Jethava is Associate Professor and Director of the Multiple Myeloma program as well as Associate Director of the Blood and Marrow Program at the University of Iowa. Prior to this appointment, he was director of the Bone Marrow and Stem Cell Transplant Services Division of Hematology Oncology at the Winthrop P. Rockefeller Cancer Institute at University of Arkansas for Medical Sciences or UAMS where he established the first fully functional allo transplant unit and he was Director and Attending Physician for the Leukemia and Lymphoma Subspecialty Clinic there.  

Dr. Jethava is an International Myeloma Working Group member and part of the EMBT malignancy working group and is a member of the Experimental Therapeutics groups and others. He reviews abstracts for publications like blood leukemia in the Blood Cancer Journal, and he is also working on studies that involves high dose vitamin C infusions for plasma cell disorders like myeloma and expects a trial to open soon at the University of Iowa. Thank you to both of you for joining us. We're thrilled to be able to fund some of your research on these important topics. Let's get started with some of the basics, like maybe the most basic question is what are myeloma stem cells? 

Dr. Zhan: Myeloma stem cells are similar to other cancer stem cells. Sometimes they are also called tumor initiation cells. Recently in a document described in Leukemia they were described as a rare population of cells with unlimited self-renewal capabilities. Renewal of tumor-initiating cells or myeloma stem cell have identified tumor-initiating cells in other hematologic and solid tumors. These cells have drug resistance or chemotherapy resistance. Recurrence of cancers after treatment, including multiple myeloma is most likely due to the persistence of a minor population of cancer stem cells which is non-cell cyclin or low cell cyclin and a very drug resistant myeloma population.

Jenny: Stem cells are these early precursor cells that become other types of cells in the future, right?

Dr. Zhan: Right.

Jenny: As they mature, because ultimately, some of them become plasma cells that we're familiar with in myeloma.

Dr. Jethava: Right. That is a very interesting fact you pointed out, Jenny. But that is a different type of stem cell that you are mentioning. It's a hematopoietic stem cell. In our blood, there are red cells, white cells, platelets and all other components, they come from hematopoietic stem cells which is the originating cells. Now, there is increasing belief that cancer also arises from one particular cell which has the capacity to keep growing and be drug resistant, and that is called cancer stem cells.

Jenny: Oh, okay. I was not familiar with that.

Dr. Jethava: What you mentioned, that's the hematopoietic stem cell. That's a completely different thing. “Stem cell” is the feature of renewability, continuous renewability and we now believe that lots of hematological cancer and solid tumors arise from a cell which is an originating cell like a tumor-initiating cell or cancer stem cell as Dr. Zhan mentioned.

Jenny: Dr. Zhan, you mentioned that they were resistant to chemotherapy, but can you go into a little bit more about or both of you about why they're chemo resistant, what makes them different from these other types of stem cells?

Dr. Zhan: Right. First, this sort of myeloma stem cells have a special expression network. As you mentioned earlier that induced pluripotent stem cells or embryonic stem cells, those cells are special expression genes such as Oct4, Sox2, and Nanog. Myeloma stem cells also express these genes. These genes drive myeloma cells self-renewal, that's one major reason. Secondly, efflux drug pump proteins are highly expressed in myeloma stem cells. Those proteins pump the chemotherapy drug outside of the cancer cells. In other words, these drugs cannot stay in in the tumor cells for a longer time, just a very short time. That's why they cannot play in an effective way to cure tumor cells. I think it's the major reason.

Jenny: Because the drug can't stay in the tumor cell is what you're saying?

Dr. Zhan: Right.

Jenny: So it's not as likely to be able to kill it?

Dr. Jethava: Correct. As Dr. Zhan mentioned, see, there are certain reason why we call cells as “stem cells”. The first and important thing is that they have higher expression of genes which Dr. Zhan mentioned. These genes are responsible for self-renewal, so they keep replicating. It means that the disease persists. The second reason is that they also have some proteins on their surface which basically pump out the chemotherapy drug out of the cells. That way, the cells become resistant to the chemotherapy. It's like a double whammy. There is this pump which is pushing out all the chemotherapy from the cell into outside the cell. So the cells are not effectively killed, plus they have this higher expression of genes which will keep them replicating. They are chemo resistant and they keep replicating. That is the reason why these cells persist for a longer duration.

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Jenny: Okay, sorry. I'm taking notes.

Dr. Jethava: No problem.

Jenny: Yes, that makes a lot of sense that they have this double hit basically. That's why they're so resistant to chemotherapy cells. Does everybody have these cancer stem cells in them and then some people, their immune system is just able to kill it or is this just something unique to myeloma where these cells just generate somehow and then they keep proliferating like you were saying and then they don't respond to treatment -- in the general population, do these types of stem cells exist I guess is my question?

Dr. Zhan: If they are debating about myeloma stem cells, some people believe that they're a small population like cancer stem cells. They can self-renew and are drug resistant. Some physicians or scientists, they don't believe that myeloma stem cells exist in this disease. However, based on our almost 20 years' experience especially for using Gene Expression Profiling, we found some patients even in the longer remission such as Total Therapy 1 patients treated at Little Rock UAMS (University of Arkansas). Those patients we can see survival over 10 years or 15 years, we collected those sample and did the gene expression profiling. We found that those “Spike” expression genes indicating translocation genetic changes exist on those for complete remission samples.

That suggests that a rare population of myeloma cells exist even in the longer complete remission samples. From that way, I started our projects that we did with Dr. Guido Tricot when we moved to the University of Utah almost ten years ago. We started to identify or define drug resistant cells or signaling pathways on myeloma stem cells. Since then, we have been in the last five to six years identified phenotypic markers of myeloma stem cells  in this project, the CD24 positive myeloma cells showed increased drug resistance and self-renewal and the sort of myeloma stem cell features. Coming back to your question, I believe most of cancers I think have the universal cancer stem cells. Probably, that's the major reason for the disease reoccurrence or relapse.

Dr. Jethava: Coming to your question, Jenny. Your question was that do everybody -- every cancer has stem cells, right? That was your question.

Jenny: Yes.

Dr. Jethava: There are two philosophies here. One philosophy, and the jury is out there regarding this thing. But there is increasing evidence to suggest that cancer arises from tumor-initiating cells. We can call it tumor-initiating cells or cancer stem cells. Is there 100% evidence? Probably no, not right now. However, increasingly, people in the field believe that that is the reason why cancer occurs, that there is a tumor-initiating cell which causes cancer but that cell is very difficult to get rid of and that is what leads to persistent disease and eventual relapse of the disease.

This whole research started by Dr. Zhan and Dr. Guido Tricot from the original data that was generated on the Total Therapy trials. I don't know whether you are familiar with that terminology or not. But just to briefly explain on what Total Therapy trial is - Total Therapy means using all drugs upfront for treatment of myeloma, all available drugs up front. This was started by Dr. Bart Barlogie and Dr. Guido Tricot in Arkansas. All patients were treated in a similar fashion. They had gene expression profiling tested at regular intervals. What Dr. Zhan and Dr. Tricot found was there was certain number of patients who were in complete remission for say 10, 15 years. There had this particular phenotype of cells in their bone marrow. That phenotype persisted for a long time, and that led to subsequent research in last ten years which is now what we are seeing this concept of myeloma tumor-initiating cells. That's the summary of the whole thing.

Jenny: I think the benefit of using the data from the University of Arkansas was that you have this really long-term data when you're saying over 10 or 15 years that you're following people and you are looking at patients in remission and then you are able to test their genetics before and after and that's pretty amazing that you have that long of data to see.

Dr. Jethava: They're unique, exactly. We have patients who are going on for 15 years and 20 years, and we have their genetic data. That's what enables us to do this kind of research.  

Jenny: When you said you were looking at these patients over 10 to 15 years, it sounded to me like you were saying that you were able to see their genetic changes and that patients who were lasting that long had similar genetic features from when they started. Is that what you were saying or were you saying something else?

Dr. Zhan: For this particular question regarding the Total Therapy 1 patients, actually we don't have a sample collected at the diagnosis because when we started our gene expression profiling these the patients were already treated. This is the reason why I mentioned that patients over 10 years have an obvious complete remission. However, Total Therapy II we have some samples in the diagnostic stage and in remission in the lab which we can confirm our observations of these genetic changes.

Jenny: That makes sense. For the first group, you didn't have a starting genetic profile because maybe you didn't have a test out yet. But then the second group did. They have the before and after which I think is really important and you have done so much work on the genetics. It's been your focus. For a lot of your focus - you've done a great work there.

Dr. Zhan: Thank you. 

Jenny: So we are thankful to you both. Let's talk about relapse after therapy. If you have these tumor initiating cells or myeloma initiating cells that causes the myeloma cancer at the beginning. And then these cells are both resistant to treatment and then keep replicating.  You give the patient treatment (induction therapy, stem cell transplant, maintenance therapy) and then they still relapsed. How do you determine who might be more prone to do that to relapse or if these cells really are in each patient and how do you classify them? Anyway, is there any way of understanding who is likely to relapse when you look at things like this?

Dr. Zhan: Yes. Once we identified that this is CD24 as a specific marker related to drug resistance or “stem cell” feature, we check this marker expression in newly diagnosed patients and also patients after treatment. So from our current data, of course the number is not big yet. I think we had totally 60 samples including 28 from newly diagnosed patients, and 32 from from patients after treatment. We observed an overall CD24 positive population, the percentage is related to higher after treatment compared to some baseline data collected at a newly diagnostic stage. With these data we hypothesized that CD24 positive myeloma cells are drug resistant.

After treatment, these cells can survive and be enriched, which is why these cells can be detected relative to a high percentage after treatment or even after stem cell transplant. This population can stay for a longer time or relatively a short time, we don't know for remission depend on multiple factors that you have the intrinsic genetic change factors. Also you have the microenvironment like the bone marrow niche of immune response make this population to grow faster and get a disease relapse. We tried to say, for any way we can depend on this phenotype CD24 population to predict the patient: the longer or shorter in remission, then the faster or slower for relapse.

Jenny: When we talk about patients, I think some patients might not understand what some what some of these CD things are. Can you explain maybe how you picked up in the flow cytometry test -- because most patients will go and get their flow cytometry test run. It'll come back with things like oh, you have presence of CD56 or you have presence of CD138 or things like that. I think they're probably more familiar with some of those even though most of us have no idea what they mean for our myeloma.

Dr. Zhan: That's right.

Jenny: But this is one of those other markers, right, that you can find on the surface of myeloma cells?

Dr. Zhan: That's right.

Dr. Jethava: Correct. What CD is basically every cell in our body has a unique CD marker on the top of it, because of which, we are able to recognize the cells. Myeloma cell has certain markers on the surface which tells us that they are myeloma cells. One of them is CD38 which you mentioned, CD138, CD19. These are all myeloma cell markers. Now, on the surface of this myeloma cells, there is this another marker also which is CD24 and this is what we are interested in. This test is called flow cytometry which is a part of bone marrow examination. Whenever a patient gets bone marrow done, there are certain things we do. One is we see the bone marrow under the microscope and see how many, what is the percentage of cancer cells. The second thing we do is do a flow cytometry, and that means we run this bone marrow sample through a machine which will pick up the CD markers and will tell us that okay, this patient has the CD38, CD138 positivity which basically tells us more specific details about the cancer cells.

And then lastly, we also do something called FISH testing which is basically to see the chromosomal rearrangement on the cancer cells, in the cancer cells. These are the three standard tests done for any bone marrow. CD markers are usually picked up by flow cytometry and CD24 is one of these markers which we are interested in. As Dr. Zhan mentioned that what our initial data shows -- I mean this is what we are planning to do like see the CD marker in the beginning of the treatment, after the treatment, how many cells, CD24 positive cells persist, what is the correlation with the relapse. That's our hypothesis.

Jenny: Let me ask you this because I think sometimes you look at genetics and they affect certain proteins on the service of myeloma cells. Patients have heard a lot about things like BCMA and CS1 because elotuzumab is going after that. These CD markers aren't necessarily protein targets or how are they -- they're just what you just explained. They're just markers but they are targets too, right?

Dr. Zhan: Right. This CD24 marker is relatively easy to be targeted because scientists can develop an antibody or CAR-T to target those specific markers easily for the cell surface genes or proteins. That's why the current CAR-T like the CAR-T19 or BCMA, that's all cell surface markers and the same thing – our goal (if we prove that CD24 is a myeloma stem cell marker) is to try to do is develop a specific antibody to target this specific CD24 biomarker – it’s a signaling pathway, or combine it with a CAR-T or develop a CD24 bi-specific antibody. Specifically, you recognize myeloma cells CD138 and also CD24. Yes, that's our goal. We will try to do it in this way.

Jenny: In proteins, when you talk about myeloma genetics and you say gene deletion 17 affects this TP53 gene, and that gene does something really specific, like it's the master cell regulator of death for all the other cells which is why that genetic feature is usually aggressive in myeloma. What does CD24 do, like does it have a specific function? I'm just curious about that.

Dr. Jethava: CD24 is not a gene. It is present on the cell's surface. What it does is that it gives this overexpression or more population of CD24 cells. It leads to more cell survival. What happens is that CD24 peptide which is basically present on the cell's surface, it activates multiple signaling pathways within the plasma cells, and these pathways are called MAPK, NF-kappaB, Notch, Hedgehog signaling pathway. This CD24 peptide is associated with activation of all these multiple downstream pathways which leads to active cancer cells, which leads to basically the resistance in the cancer stem cells.

Dr. Zhan: Regarding your question about the connection or correlation with a clinical or genetic change. I'm sorry. As I mentioned earlier, we only tested the 28 newly diagnosed patients. So we don't have data to test the correlation between genetic changes with the CD24 expression at this moment. If our hypothesis is correct, CD24 is only expressed in a small population of myeloma cells from the same patient. That means, even you have a translocation or without the translocation or deletion 17 maybe there is not any association between these types of  genetic changes.

However, recently we did an interesting study. When we co-cultured myeloma cells with stromal cells. We can see CD24 expression relatively increased. Also as I mentioned earlier, the drug treatment also can enrich CD24 positive myeloma cells. Of course, these CD24 positive myeloma cells can become CD24 negative cells. It's not on the change unidirectional but bidirectional and dynamically, CD24 expression change. Most of things from this environment, microenvironment of the drug treatment, can influence CD24 positive myeloma cell originality from the sample, even from the same sample. We can see difference in the CD24 expressions even in the same genetic background.

Jenny: That's so interesting. In your initial study, you said you took the 60 samples and 28 were newly diagnosed and 32 were relapsed patients. And you were just looking for the levels of that CD24 expression to see. That's so interesting that you just said that if the application of drugs, some drugs can actually increase the level of CD24. 

Dr. Zhan: Right. That's a two-way. One is probably directly up-regulated CD24 expression. Another one is the CD24 negative cells are killed. Only CD24 positive cells can survive, so it really enriches the CD24 positive cells. We believe with the later one, enriched CD24 positive cells, the more likely the upregulation of CD24 expression, but we needed more study to confirm this observation.

Jenny: Overall, I mean you're looking at these samples to say -- and your hypothesis that just higher levels of CD24 expression means tougher disease or more likelihood to relapse I guess. Is that correct?

Dr. Zhan: Yes. The short answer is yes. We think the high percentage of CD24 positive myeloma cells are relatively insensitive or resistant to current  treatment therapy or even transplant. That these cells can survive and can initiate the disease relapse. That's our bigger hypothesis.

Jenny: Right. That's what's causing the relapse because these cells are still sticking around and they're still tumor-initiating and they're still replicating and they're not being killed by the drugs that you're getting, right?

Dr. Zhan: That's right.

Dr. Jethava: Correct. To give you one example, Jenny. Let's say you have a patient, patient gets treatment with standard of care which is Velcade, Revlimid, dexamethasone. They get treatment after four cycles. You see that they have not achieved adequate response. In that case, it would be worth looking at why they are not achieving response. Because typically what happens is that patients, they receive second line of treatment which is usually Kyprolis/Rev/Dex or Kyprolis pomalidomide, dexamethasone or they go for transplant or they go for daratumumab combinations. In spite of all these combinations, certain percentage of patients don't achieve MRD negativity and they keep relapsing. In those patients, we believe that there will be increasing proportion of CD24 cells within the plasma cells, within the monoclonal plasma cells which is what is keeping the persistent disease which is what is leading to the relapse. 

Jenny: Right. What you mentioned is that the first part of your study and one of the goals of your project is to understand how much CD24 does a patient have. Are you separating that out again between newly diagnosed and relapsed patients? Are you just going to look at relapsed patients, or who are you going to be testing?

Dr. Zhan: In this proposal to the Myeloma Crowd Research Initiative, we proposed to test 60 newly diagnosed patients. But we will also do the follow-up or collect the sample at different  stages like we collect the sample at diagnosis, in remission after treatment and also a sample at the relapse. We can follow-up this series of samples – do they change? I'll see if this really can be used as a marker to predict the patients who may experience early relapse or become resistant. That's our goal.

Jenny: You're going to be using samples from existing University of Iowa patients, correct?

Dr. Zhan: In this moment, yes, the reason is we have intellectual property in process. Once we get this approved, I think we can open it up soon. Another thing is we have a manuscript under review in the Journal of NCI. Once these are published then the information is available to the public.  

Dr. Jethava: Right now, we'll be doing it at University of Iowa, this testing. Then as Dr. Zhan mentioned, there is some intellectual property thing going on with it. We are hoping that it will be all cleared and then the revision of the submitted manuscript is getting ready. We are hoping that in the next few months, it will be ready for everybody.

Jenny: Great. And then the second part of your study is to say, if we have a lot of CD24 in a particular patient, then what drug targets could go after that target as far as a monoclonal antibody, right?

Dr. Jethava: Right. That's exactly what the plan is.  Let's say we see a patient who is at high risk as you mentioned, say bi-allelic 17p deletion and then patient has not achieved MRD negativity and then we see that they have increasing portion of CD24 in the monoclonal plasma cells. Rather than just blindly going for salvage transplant or some kind of newer modalities, the best thing will be to target specifically the surface CD24 with a bi-specific antibody or by using some other technique. Dr. Zhan is working on that as well.

Dr. Zhan: That's right.

Jenny: I think this idea of identifying the cause so you can impact the effect is really needed in myeloma because right now, when people relapse, well, they look at the genetics for sure and that does have an impact but there are other things that are potentially part of that whole picture. The more you can personalize it and say oh, we do see higher levels of the CD24 and then let's go after it with this particular drug, that would be fantastic.

Dr. Zhan: Right. That's correct. Now, actually, we are collaborating with a professor in Washington U, Dr. John DiPersio to develop a CD24 CAR T and they are also trying to develop a bi-specific antibody that should have both CD138 and the CD24 positive antibody. We are doing this work, but then we need some time to work this out.

Jenny: Right. One of my questions was going to be - are there any CD24 drugs that already exist or antibodies that exist, but it sounds like you're going after myeloma partnership to develop that and then test it?

Dr. Zhan: Right. There is one paper published from a Chinese group. They published in the Oncotargetjournal to engineer a high-affinity humanized anti-CD24 antibody to target the hepatocellular carcinoma liver cancer. It's been done in a pre-clinical mouse model.

Currently, I don't think they have done this in a human sample or cancer patient yet. However, because CD24 is a ligand for P-selectin,  The anti P selecting is involved in treating sickle cell anemia. I'm thinking to blocking this binding between this CD24  with p-selectin like anti P-selectin antibody maybe can be coincided for using -- in treating cancer. That's one we needed to do, but we didn't start this experiment yet.

Jenny: Well, that's really neat that you're looking at existing things that are being used in other diseases or other cancers and then being able to bring them to myeloma. That's more efficient than trying to create your own.  But at WashU, they're both creating a CAR-T cell going after CD24 and the bi-specific antibody? Is that what I heard you saying?

Dr. Zhan: Yes. We had a discussion about three months ago. They tried to develop it both in both ways.

Jenny: Interesting. That was so fascinating. Are they seeing other uses for that CD24 or just in myeloma?

Dr. Zhan: In this stage we can start with myeloma because we don't know clearly CD24 expression in other cancers of course. Like breast cancer, people thought the CD24 is negative or  not expressed in the stem cells. However, with the positive CD24 cancer is a marker for liver cancer stem cells. Dr. DiPersio, he is also a hematologist. In this one, we think more is in myeloma but that depends on future if we test the CD24 could be a marker for other cancers or other disease. We need to think more in the future.

Jenny: When you think about this, the CD24, let's say you developed this CAR-T cell or this bi-specific antibody or something like that but can go after that target. How could you envision it being used in a clinic? Would you give it upfront as part of your induction therapy so that the cancer cells, you're hitting certain cancer cells when you give a transplant or where you give RVD or something like that. You know you're hitting a lot but maybe not all of it. Do you just add it at the front end or do you get through that part until when the tumor burden is lower and then you just say okay, now, we're going to give you a CD24 thing to kill off these stem cells that are proliferating?

Dr. Jethava: That's a very good question, Jenny. This is an evolving strategy. This is a very good question you asked. It makes sense to use it upfront after the induction chemotherapy when there is minimum residual disease left and target it with CD24 specific antibody either bi-specific or a CAR-T so that we try to wipe out this tumor initiating cells upfront. That's the strategy we are thinking about.

Dr. Zhan: Currently, there are a lot treatments that can induce remission in patients but the problem is the patient relapses after a while in remission. Our goal is to develop a treatment that can prevent the disease relapse. Directly answering your question is we think this treatment should be when the tumor burden is low after care, then it's a good time to add this specific treatment to drug resistant myeloma stem cells.

Jenny: Well, that makes sense. You kill most of everything and then when the tumor burden is low, you go after that. I had a question going back to what you're saying before. This is a flow cytometry test. Forgive me because I can't remember. That's just a blood test, right?

Dr. Jethava: No. that's a bone marrow test.

Jenny: That's a bone marrow test, okay. Does every flow cytometry test currently look at those CD24 levels or is that something that you have to especially request or the lab has to run?

Dr. Jethava: First of all, when we do the flow cytometry, there are typical markers looked at.  CD24 is not one of them. The individual lab has to standardize the test and then validate it and then run it. Like say someone is in Kansas, University of Kansas and they request of their hematologist, can you can please check CD24 fraction on myeloma cells? They might not be able to do it immediately because they need validation making sure that the test is replicable and is working well before they can start giving the results out. These are the technical issues with the test. In our institute, we have overcome those issues right now, so we are able to do it routinely on the patients here at the University of Iowa.

Jenny: Well, that makes sense. I think this could influence testing and treatment in the future if you find that this is true and that you have a target that can go after it. I think people would start changing their test to be able to predict for that because then you have more personalized medicine.

Dr. Jethava: Right. This is basically a really personalized medicine where we are going after one particular marker to see how the things will pan out in the future. We are very excited about it.

Jenny: I think it's wonderful. I know there are other doctors too that are trying to go after the stem cells. Even in the CAR-T environment, most patients don't have a lot of CD19 on their myeloma cells. That one patient that had such a dramatic response with one of the first CAR-T treatments at UPENN, they're really surprised that she had that response, but I remember them saying, "Oh, well these might be some of these early precursor cells." So trying to find out why people are relapsing and going after it is truly amazing because a lot of patients have really good responses with the treatments. But like you said, over time they relapse. If you could knock everything down and get rid of it all, that's really amazing.

Dr. Zhan: Right. The CD19 and CD24 both are expressed in different stages. However, they are not simultaneously expressed in the same stage. Sometimes, they express CD19, sometimes CD24 or both. Currently, we tested eight patients when I worked with Dr Salama, at the University of Utah. He found that three of eight patients had both CD19 and CD24 expression.

Jenny: Oh, interesting.

Dr. Zhan: There another five that only expressed CD24, and no CD19 expression. But that sample size is too small, we needed to do more in the future.

Jenny: Yes, that's so amazing because you could get to a point where you just do it individually for each patient and say well, you're going to need this drug combination and you're going to need this drug combination. That's I think so promising in myeloma and so exciting in myeloma that you are getting to this point. It's really amazing. How long do you predict the study will take and how long will you need to watch the patients? Just curious.

Dr. Zhan: Right. We found this in clinical samples but using the flow cytometry to detect CD24, we are almost at one year studying this marker, around the one year, right?

Dr. Jethava: Correct, yes. Correct.

Dr. Zhan: So far, we don't have a high clinical outcome yet because one year is too short to say a good prognosis or bad prognosis. We have two goals: One is the prognostic marker or drug response correlating with levels of CD24. Another one is, if this correct, we need to develop a treatment targeting CD24. I think in the next probably three to five years, there is one group in my lab who are focused on this project that I think we need at this time.

Jenny: Well that's great. And then we talked earlier in the show about how these projects will integrate with HealthTree. How do you envision this integrating with HealthTree?

Dr. Jethava: In my clinic, we have started asking patients to register themselves on HealthTree. They will be putting their own information. And then as a part of HealthTree initiative, we have discussed about including this particular marker on their flow cytometry testing list or in one of the sections like to some kind of bar can be created. And then patients can ask what is my CD24 number and doctors can tell them that this is the number. They can just go ahead and add it in there. In that way, there is running data available. There is always updated data available for these patients. That's the plan to integrate it with the HealthTree.

This will help in numerous ways. The first way, it will be patients -- say if there are 500 patients with CD24 number newly diagnosed. What's happening with the treatment? How many are responding? What's happening to the CD24 number after their response? All this can be looked at with a more detail and with a broader view. That's what we think we should be doing with the HealthTree initiative and to integrate it with the HealthTree.

Jenny: With HealthTree, you'll also be able to include CD24 and then you can also add a percentage. This happens too on genetic deletions and things like that. Not every patient has the same percentage. Some patients might have 5% of CD24 and some patients might have 80% of CD24. That percentage is kind of important, right?

Dr. Jethava: Correct. That percentage is important as well. HealthTree will be anonymously beneficial for seeing the data. For example, we have 500 patients, we have their CD24 number. Out of that 50 patients don't respond to the treatment. What was their cytogenetic FISH risk and how did that correlate? Was there any correlation or there wasn't any correlation? All questions can be answered with the HealthTree data.

Jenny: I think it's really important. Well, we encourage patients who are listening to join HealthTree because this is a way that we can help the research community get to these types of conclusions faster. Now, I want to leave just a few minutes for maybe one or so questions because we're running short on time. But if you have a question for Dr. Zhan and Dr. Jethava, you can call 347-637-2631 and press 1 on your keypad. Our first question go ahead with your question.

Caller: Hi. Thank you for having me on. Dr. Zhan mentioned that there is a debate about the existence of myeloma stem cells. I'm just wondering why there is a debate at all and why can't we just easily see whether they exist or not?

Dr. Jethava: First of all, it's very difficult to diagnose. What is basically a feature of stemness? The feature of stemness is that these are the cells which persist for a long period of time and they are responsible for relapse. To characterize these cells, we need to have patients who have lived more than 10 years or 15 years or those who have had relapses. We need to find this population in this particular cell population in group of patients who have been in remission for a long time to see what is the characteristic of these cells. It's very difficult to find these patients who have lived for so long. Plus, as Dr. Zhan mentioned initially, gene expression profiling and all these modern techniques were not available until 20 years ago. Now, they are more and more available. There are technical issues and there are logistical issues which has led to difficulty of characterizing these cells.

Dr. Zhan: When we see the cell lines in culture in vitro, 99% of cells are CD24 negative. However, these cells can divide and then propagate the proliferation. That's one concept. There are currently two concepts are for cancer stem cells. One is that every cancer cell could be cancer stem cell. Another one is like we proposed that only a small population of tumor cells have stem cell features. That's the reason some people believe the first concept, some people believe the second one. From currently our data, when we collect CD24 positive cells from myeloma cell lines, we find that only 10 cells can generate a tumor in mice. However, when we use the CD24 negative cells, you can gave 10,000 cells and no tumor formulation. That's big evidence that validates the second concept. Only a small population have stem cell features. I hope this is understandable.  

Caller: Yes, a little better. Thank you.

Dr. Jethava: What Dr. Zhan said, I'll probably repeat it. There are two concepts, two philosophies. One is that small population of stem cells lead to the cancer relapse and progression. The second philosophy is that every cell, every cancer cell is a stem cell. We believe in the first philosophy that small percentages of cells lead to cancer generation, and that is based on our laboratory experiment. What Dr. Zhan's lab has shown that if you take CD24 positive cell line from myeloma patients, cell line, only 10 cells are enough to produce myeloma. That's it. Now, if you take CD24 negative cells and even if you inject 10,000 cells in mice, it will not lead to the myeloma formation. This is the reason why we believe that even such a tiny number like stem cell can lead to myeloma formation, we believe that this might be the marker. That's just right now our hypothesis.

Jenny: That's amazing. Okay, thank you for your answers and your question. One more question, go ahead with your question.

Caller: Hi, Dr. Zhan. You mentioned that you had been tracking some patients at the University of Utah with the CD24 present. Have you been able to track the outcomes of those patients and how they responded to treatment?

Dr. Zhan: Thank you for this question.  Actually, short answer is no because this is not done by myself. There is a collaborator, a pathologist, Dr. Salama, who is now at the Mayo Clinic in Minnesota. Also the patient information was de-identified information that only included eight samples. We really didn't think we should follow-up on patient outcomes because it's a smaller sample size, but in the future we have to do it. We should do that.

Dr. Jethava: Right. At the University of Iowa now, we have identified the patients who have MRD negativity and who have CD24 fraction positive in them. We are following them. I cannot disclose everything right now, but we have sizeable number of patients we are following closely and we are monitoring their CD24 on bone marrow and it's interesting because some of them -- these patients have no disease biochemically. Biochemically, there is no M-protein. They have normal kappa/lambda ratio. But then, when you do the bone marrow, and we have a very sophisticated MRD detection which is by the next generation flow. We do it here and by next generation flow. We see that if they are MRD positive and they have CD24 fraction, then we are closely following them to see what's going to happen. It's evolving right now.

Caller: This is exciting yet maddening because most of us, myeloma caregivers and patients, do not care about being hidden in the de-identified database. In fact, it's not really helping us. That information for those eight de-identified patients is going to be forever lost somewhere and you have in there some potential clues for hypothesis that strengthens what you guys are doing. I think this is really interesting in terms of how this does intersect with HealthTree and how this could really accelerate this type of research and protect the patient's information, but yet not lose it in some obscure IRB study.

Dr. Jethava: Well, no. We totally appreciate your point. I guess the issue is that when do we want to inform the community, right? I mean we don't want to be going out and telling the information which we ourselves are not sure. I think when Dr. Zhan and this was done in Utah, at the time, the patient number was small that it was probably not the best thing to do anything with that. I think in last seven, eight years, the research has become mature and more information is available. Now, I think in going forward, it will be very important to track this information.

Dr. Zhan: Right. That's certainly it.

Caller: Well, thank you.

Dr. Jethava: Thank you.

Dr. Zhan: Thank you.

Jenny: Thank you for your question.  Well, Dr. Zhan and Dr. Jethava, we're so appreciative for you, for the research that you're doing and for joining us today to help explain it and we're very, very excited about what you're doing. It's clear because we selected you to receive a grant and to go after this for us. I think this has been a mystery for a long time and I am grateful that you have such key experience working on it, and we're just so thankful that you're looking at this and studying it on our behalf.

Dr. Zhan: Thank you all very much. Thank you, Jenny, Yogesh. Good day.

Dr. Jethava: Thank you very much, Jenny, and we appreciate your help. Thank you.

Jenny: Well, thank you so much and thank you so much for our listeners for listening to Myeloma Crowd Radio.  We invite you next time to learn more about the latest in myeloma research and what it means for you.