Questions and Answers Video (Text Version)
Title of Talk: Question and Answer Session
Speaker
Thank you Dr. Vessella. We’re going to open the floor for questions.
Speaker
Gail Prince, the University of Illinois. I have a question for Marianne. So in the interaction of your EPI001 molecule with the N-terminal domain; so do you know which amino acids it’s associating with? You showed the region where there’s a conformational change that’s essential. But do you know what sites it’s actually ligand(ing) to and with the possibility of explaining how a conformational change could occur?
Marianne Sadar
No; that’s–that’s a wonderful question. We–we are currently working with some mass spec facilities to try to identify what–what that amino acid or those amino acids are. We’re also trying to approach this by NMR and so far we don’t have any data on that. I’ll expand though; so I showed you that Mutation 8 that Ian McKeown–he’s done the work on that. He also mutated some amino acids in some of the other alpha-helical regions and we got the same data that–that destroying those regions too completely destroyed the interaction.
Speaker
Okay; because that could be key to identifying you know future–third and fourth generation products there, so–.
Marianne Sadar
Yeah; so–so we don’t know those amino acids. We do know that we don’t inhibit all protein interactions. I showed you data yesterday about CBP; in the paper we also have RAP74. When we looked at the steroid receptor co-activators, we couldn’t–well I shouldn’t say we couldn’t–we still detected interaction of the steroid receptor co-activators to the N-terminus and these–these co-activators can also act as repressors in certain situations, so it doesn’t inhibit everything.
Speaker
Okay; thank you.
Speaker
I actually wanted to ask two questions; is that possible?
Speaker
If they’re quick. We do have some time.
Speaker
Very quick–so I’m going to begin with the one that I think has probably a difficult answer to and I’m going to forward that to Dr. Sadar and then the other one to Dr. Vessella. I want to congratulate both of you on a fantastic presentation. So this is a clinical observation; to date there has not been a single example where therapy directed at the receptor directly has eclipsed suppression of production. So if you think of it in terms of the orchiectomy or the LHRH therapy you have not been–we have not been able to show that a receptor blockade eclipses that. And certainly in the context of say the Abiraterone it is a production example, again showing that the minute you reduce production it makes a difference. And if the Medivation drug makes it actually–so that’s sort of the son of Bicalutamide so to speak, you’re talking about in potential mutated or altered receptor and even that is going to come at the backbone of already suppression of production that you’re doing by either an orch or an LHRH. Any type of I guess thinking or thoughts as to what is it about the receptor that we cannot really block it well? And so that would be the one question.
And then the question to Bob Vessella who did a fantastic job and I want to echo your request of looking at the mets to design treatment, not the primary because it’s a no-brainer; the disease is totally different–why is that with all the technology we have now even in the setting of establishment of bone mets, we only actually pick up 50% with circulating cells or disseminated cells? Thank you.
Marianne Sadar
So with–with the current therapies that target androgen receptor in the ligand binding domain, there’s speculation that–that these compounds fail. I mean it will be good when we actually find out definitively what the mechanisms of resistance are for Abiraterone and for the MDV drug. I can only speculate that–that right now it’s because of the splice variants. There is some data starting to support that. The more you decrease the tumor levels of androgens the more highly expressed some of these variants become. And that’s really some work that’s been done at the University of Washington. So these variants may play one of the key roles in these resistant mechanisms. There also may be slippery ways to get around SIP17 that–that other steroids may be–becoming up-regulated. So the–the hopes that an N-terminal inhibitor could circumvent that–the hopes are quite high right now. You can get potentially mutations; I showed you potential mutations in the alpha-helical regions that perhaps those would happen but if that happens the receptor shouldn’t be active anyways. So an intrinsically disordered region, mutations have a less chance of actually doing much. So the N-terminus domain, I–it’s attractive on paper. Whether that pans out in the clinic, we’ll have to wait and see.
Robert Vessella
And [Mahat] were you asking why we detect 50% in the–of disseminated tumor cells in these advanced staged patients? Was that–was that your question?
Speaker
So in the setting of metastatic disease, you should detect 100%, right?
Robert Vessella
Yes.
Speaker
Right; because it’s already disseminated. And so is it because there is a lousy technology which is what we have today that we are unable to detect these cells or is it because somehow the cells get smarter and they evade detection?
Robert Vessella
I think–I think that–I think both are contributing. First of all, we only–when we do the bone marrow aspirate we’re only taking from the ileac crest, so there’s certainly a sampling error. If I had the ability to take multiple bone marrow aspirates, I’m sure we would have 100% in all the patients because as I said shortly after taking those bone marrow aspirates if these patients undergo a rapid autopsy, we can see that 70% of the bone sites do have disease. So you’re absolutely right; whether they’re circulating or in the bone marrow, the cells should be there.
Early on, we also have the problem of this epithelial to mesenchymal transition. Now EPCAM we have found to be the more durable of the various epithelial markers. But I would certainly predict that we’re missing some tumor cells that have also lost EPCAM and they’ve lost PSMA, they’ve lost EPCAM, maybe cytokeratin and so they’re still there; they’re still a tumor cell but we don’t have a handle on which to capture them. So I mean if I were to go out on a limb, a pretty far limb, based on the experience that I’ve had so far in looking at these little buggers for close to 15 years I think every patient–the first time they come into the clinic, the cells have escaped. They have circulating tumor cells; they have disseminated tumor cells. Some are quite a bit more virulent or aggressive than the others, but if you say is it a tumor cell–did that patient have a-tumor cell of whatever phenotype in their circulation I would say yes, 100%. The cells have–have escaped. Can we detect 100%? Not with the technology that we have available today.
Speaker
I think in the back was the next question.
Speaker
My question for Bob–my question is for Bob Vessella. Bob, in the last couple slides you showed that in a given patient with multiple lesions that you could have anything from PSA positive to PSA negative and AR positive and AR negative. Have you looked to see if there is neuro-endocrine differentiation occurring looking for like chromogranin-A in these samples?
Robert Vessella
We’ve–we’ve–yeah; we’ve looked at chromogranin-A and synaptophysin and a variety of others. Those are–we don’t often see a high expression of those markers. There’s a heterogeneity in the expression but it’s usually at a very low percent, so that one site might have 10 or 15% chromogranin-A positive and another site might have 1% or whatever. But you don’t often see or at least we haven't seen in our series where one site is 100% and another site is zero. It fluctuates probably 15% or below.
Speaker
So you’re saying that the level is low within a given site but if you looked at 10 sites would all 10 be positive or would one site be positive?
Robert Vessella
Well again depending on what–what do you call 1% positive?
Speaker
I would call any detection of synaptophysin or chromogranin-A because neuric differentiation usually occurs in isolated pockets and not totally valid cells.
Robert Vessella
Then I would say that we can detect you know 1% or so of the cells positive in most lesions.
Speaker
Okay; thank you.
Speaker
Back to the front.
Speaker
Yes. Is this on? Okay; Laurie McCall at University of Michigan. This question is also for Bob Vessella. Bob, I’m always impressed with the robustness of your data and the progress you’ve made in this area. Thank you–that there’s changes in these cells is very clear; you also mentioned the micro-environment and I think that can't be underestimated and I’m wondering if you’ve taken advantage of an opportunity to look at alterations in the micro-environment that may be reflected in association with DTCs and their progression, you know things such as bone turnover markers, you know profiles of the patients that may reflect changes in the bone marrow and how conducive those might be to the DTCs.
Robert Vessella
Yes; we have a separate project, somewhat independent of the DTC project and I showed one slide on the comparison of soft tissue mets to bone mets and then going beyond that we have a project looking within bone mets at those that are more osteoblastic with those that are osteolytic. And clearly you see gene expression differences between those two populations of cells. I absolutely concur that the micro-environment is extremely important and may in fact be the driving force in this tumor cell dormancy. As I said unfortunately we don’t know much about dormancy; we don’t know what initiates it, what maintains it, or what breaks it. We don’t even have a marker that I can use if I isolate one of these DTCs; I have no way of knowing whether it’s a dormant tumor cell. I don’t have a marker for dormancy. I know it’s not dividing or at least it’s not dividing very fast, but I do believe this concept of the vicious cycle where the micro-environment is extremely important plays a significant role and that cytokines being reduced–being produced by the–within this micro-environment will strongly influence whether tumor cells remain dormant or break dormancy.
I don’t know whether the micro-environment–one of my first slides–is involved in attracting cells to the bone, whether it’s like a big magnet that attracts cells to the bone. I’m not personally a big believer of that concept. But I do believe the micro-environment once the tumor cells get to the micro-environment that the stromal cell–tumor cell interaction is critically important. And we do have some studies under way right now using nano-technology and micro-fluidic chambers where we’re placing these DTC in these micro-fluidic chambers preceded with bone marrow stromal cells to see the differences between DTC interaction with one type of stromal cell versus another type of stromal cell. But that was a project that had just begun.
Speaker
Do–do you have measures of bone mass in these patients and/or bone turnover markers?
Robert Vessella
Not in all of the patients; in a number of the patients we do. And the–all the patients basically who have gone through the rapid autopsy program–
Speaker
I mean way before they have overt bone lesions, so when they’re still DTCs but not overtly having metastatic lesions.
Robert Vessella
They’re–they’re–did you say their weight?
Speaker
No; the bone like measures of bone mass and/or bone turnover markers that would reflect changes–?
Robert Vessella
Not–not in the early–not in the early disease but in most of the advanced disease we do but we haven't done the correlation with that.
Speaker
Okay; thank you.
Speaker
I think we have time for maybe one–two fast questions–front and then back. And then Carolyn has an announcement I can't forget to let you know about that.
Speaker
Jim [Maldenfelz], Virginia Prostate Cancer Coalition, survivor; this is a question for Dr. Thompson. In one of your slides it looked like the GLIPR-1 was only effective for one cell line. Is it more generalizable or are you aiming at a limited sub-set of the disease?
Timothy Thompson
Well I think you’re probably talking about the in vivo data, the–the protein. Now we’ve used a couple of different models, PC3 and V-CAP, orthotopic models and it’s effective in both of those model systems. Beyond that we haven't really looked at other–other in vivo systems. In vitro it’s effective in multiple prostate cancer cell lines.
Speaker
Thank you.
Speaker
And then the next one was in the back of the room.
Speaker
My question is to Dr. Sadar. And I was wondering I know you looked at apoptosis and proliferation with EPI but did you look at angiogenesis by any chance?
Marianne Sadar
Yes; I mean this–this very profound physical change in the appearance of the tumors when we treat with the sintokamides or with EPI, we have looked at angiogenesis; we’ve–we’ve done it using the CAM assay and we’ve also looked at secretion of VEGF from the tumor cells in the presence of EPI. And I can say yes; it–it does have an effect on–definitely it inhibits VEGF secretion and it also inhibits the production of blood vessels in–in the chick embryos.
Speaker
Okay one more question and that’ll be the end of questions, sorry.
Speaker
Evelyn Barrack, Henry Ford Hospital–question for Dr. Vessella. Do you have information about what percent of patients with DTCs develop clinically evident metastases?
Robert Vessella
What percent of those–well again, I can tell you in our–in our particular series about 20% of the patients post-radical prostatectomy develop overt metastases but the detection of DTCs you can go by the number that we have, which let’s say 50%, but as I mentioned a few minutes ago I would–I would think that 100% of those patients actually do have DTC but we’re not detecting them because of sampling error. But 20% of the patients post-radical prostatectomy develop metastases and a lot more than 20% have DTC.
Speaker
So the question becomes, the reason I ask the question is what percentage of patients with DTCs actually don’t develop metastases?
Robert Vessella
Well again I can tell you the vast–the majority of the patients with DTCs do not develop metastases.
Speaker
That’s good news.
Robert Vessella
Yeah; and those are probably the patients that undergo tumor cell–or that have dormant cells in their bone marrow.
Speaker
So I think we’re going to end and please see the speakers if you–afterwards outside–we have the break. I don’t want to get into the next session but I do want to congratulate all the speakers and thank them tremendously for a fantastic session. And Carolyn has the floor.