Christina Sumners: Welcome to Science Sound Off. I’m Christina Sumners.
Tim Schnettler: And I’m her co-host Tim Schnettler.
Christina Sumners: And here with us in the studio today is Dr. Rod Dashwood from the Texas A&M Institute of Biosciences and Technology. And calling in from Houston where the institute is located, we have Dr. Rajendran. Welcome to the show, both of you.
Dr. Rod Dashwood: Thank you.
Dr. Praveen Rajendran: Thank you for having me.
Christina Sumners: So we are going to talk today about the research the two of you have been doing. Just before we do, maybe just could you give our listeners a little background about yourself?
Dr. Rod Dashwood: Praveen, you want to go first?
Dr. Praveen Rajendran: Sure. My name is Praveen Rajendran. I’m a PhD in pharmacology. I’m an associate professor at the Center for Epigenetics and Disease Prevention and my research really focuses on cancer, chemoprevention and how to identify dietary factors [that] can be used for specifically for colorectal cancer and I work with colon cancer cells, preclinical models and basically focus on translational cancer research. So that’s kind of a short background about me.
Dr. Rod Dashwood: So I originally came from England. I did all of my schooling in the UK. Right after PhD, I moved to the US, 1986, as a postdoc and have spent my time in various academic institutions, Oregon State University, University of Hawaii, and now Texas A&M, with sabbaticals at the National Cancer Center in Tokyo.
Christina Sumners: Oh, wow.
Dr. Rod Dashwood: And I had a longstanding interest in how diet impacts cancer risk, both the good and bad. So what are the bad guys in food that may be causative for colorectal and liver cancer and other cancers. And in particular a great passion for dietary factors that can be used for prevention of cancer development from the earliest stages all the way through to the later stages. So I moved six years ago to Houston to join Texas A&M Health Science Center and College of Medicine. And I was brought in specifically to head up the Center for Epigenetics and Disease Prevention.
Christina Sumners: So you mentioned epigenetics. For people who don’t know what that is, could you just kind of give an overview of what epigenetics means?
Dr. Rod Dashwood: The epigenetics is “above the DNA”, so it’s really how the proteins are packaged around the DNA in such a way as to be able to fit all of that information into the nucleus, but to make the DNA still accessible at the appropriate time and place. And that often is dysregulated in cancer development. So one of the most important aspects that excites us about epigenetics is unlike the genetic changes in cancer development, which are considered irreversible. So that’s a change in DNA sequence, epigenetic changes are, at least in principle, are modifiable. So if you have an aberrant epigenetic profile in a cancer, at least in principle, you can reverse those changes through, for example, diet and lifestyle. So that’s the thinking.
Christina Sumners: Okay. So your research looks at ways that you can change the epigenetics to make people less likely to get cancer.
Dr. Rod Dashwood: That’s the ultimate goal.
Christina Sumners: Okay. Obviously that’s incredibly complicated as cancer is an incredibly complicated disease. But I know you’ve just had some recent research about colon cancer, so maybe you could explain a little bit about what y’all found out about that.
Dr. Rod Dashwood: So this is a paper that Dr. Rajendran was first author. This was in Cancer Research a few months ago, and it really is built on a long series of experiments and publications that we’ve had over the last 10 or 15 years. Before that, we were focused very much on the genetic side that I’ve described.
And it was really an accidental discovery by one of the grad students in my lab at that time that a compound in broccoli, a compound called sulforaphane, could act on the epigenetics in our cells to affect gene expression. So what we showed in cancer, and again in Cancer Research in 2004 was that this compound called sulforaphane as well as the whole food, like broccoli or broccoli sprouts, could act on the epigenetics in our cancer cells to sort of circumvent everything they’ve done to become a cancer. They were sort of opening up the chromatin, opening up access to the DNA and allowing transcription factors to gain access to, for example, tumor suppressor genes, which the cancer has shut down in order to survive as a cancer cell.
And we are tricking that system into re-expressing those genes. And now those tumor suppressor genes can be expressed and they were triggering cell cycle arrest and apoptosis. And the exciting thing that came out of that research from the very early days is it seems to be specific to cancer cells. Normal cells don’t act in the same way. So this is an exciting observation and we’ve really built on that observation over a number of publications, trying to understand how does this work in preclinical models of colorectal cancer? One of my colleagues was looking at the same system in prostate cancer. It seems to be the same, very encouragingly.
We even showed in healthy human subjects who consume one cup of broccoli sprouts that we could modify these epigenetic readouts in the circulating cells of our body. So one of the questions we often get is, “Well, how do you know that the concentrations of that compound that you’re using in your cell, in your Petri dish, or in your preclinical models have any relevance to humans?”
So our answer to that is, “Well, we know that when we eat broccoli sprouts-”
Christina Sumners: It actually changes.
Dr. Rod Dashwood: It actually changes these epigenetic endpoints. And so now we’re starting to not only look in the circulation, but also in the target tissues. So for example, we have a trial, a clinical trial with Oregon Health & Science University. People who are doing screening colonoscopies go through a very extensive cruciferous vegetable questionnaire list, over a hundred different cruciferous vegetables. You know, you could probably name cabbage, cauliflower, broccoli, maybe that’s it. Kale.
Christina Sumners: Kale, a good amount, yeah.
Dr. Rod Dashwood: This had a hundred types of cruciferous vegetables listed, serving portion size, and how it was cooked. So all that information was collected by a third party, the University of Arizona. They subdivided human subjects into high and low crucifer intake, zero to one servings a week or five or more per week.
Christina Sumners: Okay.
Dr. Rod Dashwood: And we were able to associate the reported intake in cruciferous vegetables to changes in the colon. It’s not just in the circulation but in the colon, which is our primary interest is colorectal cancer prevention.
Christina Sumners: Right.
Dr. Rod Dashwood: So that really is sort of a lot of background information got to that point. So one of the, I think key points from the paper that just came out in Cancer Research is we were really broadening our focus now beyond this… I’ve talked about the epigenetics involves proteins associated with DNA. So those are what we called histone proteins.
Christina Sumners: Okay.
Dr. Rod Dashwood: And so when people talk about epigenetics, the main things they’re talking about, number one is what’s called DNA methylation. These are like little chemical groups that are put onto the promoters of genes, which in cancer cells often silences them. And there’s the histone proteins, which kind of wrap around the DNA and are used to package it and also regulate access to the DNA.
Those can carry reversible modifications on little tails that stick out. And those modifications are what our dietary factors are influencing to open up access to the chromatin. So we’ve had a good appreciation of that mechanism, but Praveen showed through some very nice work that it’s not just the histone proteins, which are intimately connected with the DNA. There’s also other proteins that are sort of floating around in the nucleus, around the DNA—things like transcription factors, DNA repair proteins—things like that, those guys are also getting the same kind of modifications on them that the histones are. And the paper is making the point that one of those proteins, which is called CCAR2 [Cell Cycle and Apoptosis Regulator 2], that protein is actually very important for colorectal cancer development. And it goes through the same types of changes, specifically acetylation and deacetylation that are affecting its function.
So one of the key messages from this paper is, and I think a lot of people in epigenetics are really appreciating this more and more, it’s not just about the histone proteins directly in contact with the DNA, but other proteins that are playing regulatory roles in the vicinity. Their acetylation and deacetylation has an important role to play in what they’re doing.
Christina Sumners: And in whether cancer is developing or not.
Dr. Rod Dashwood: Yes.
Christina Sumners: Okay.
Dr. Praveen Rajendran: I think I basically wanted to add that we were originally looking at histone deacetylation and identifying what parts of histones are getting acetylated. But I think this particular project was focusing, like Rod mentioned, on non-histone proteins and the way we went ahead and identified some of the proteins was using a proteomics approach. So we tried to look at what at are all the different acetylated proteins that we can pick up after you treat cells with sulforaphane and that’s how we identified CCAR2 [or Cell Cycle in Apoptosis Regulator 2]. That was one of the most dominant proteins that was hyper-acetylated by sulforaphane. So that’s how we identified it. And actually our attention was drawn to CCAR2 based on I think primarily two observations. One is CCAR2 is over expressed in colorectal cancer.
Christina Sumners: Oh, okay.
Dr. Praveen Rajendran: And yeah, and there’s very nice patient data showing that patients with high levels of CCAR2 exhibit reduced survival.
Christina Sumners: Oh, okay.
Dr. Praveen Rajendran: So, which clearly shows that CCAR2 has an oncogenic role. People who have high CCAR2 in their colon, their disease is much more extensive and leads to reduced survival. So there is a need to reduce CCAR2 or its downstream signaling pathway. So the fact that CCAR2 was getting acetylated [by sulforaphane, I think] that’s where we got it. We said, “Yes, it’s important protein that’s getting acetylated, but let’s just follow it up”. And so that’s how we went ahead and carried out the research on CCAR2 in this paper.
Christina Sumners: Okay. So you saw that it was changed in cancer patients, and especially in cancer patients with poor outcomes, and so you decided to focus on that.
Dr. Rod Dashwood: So this was actually another group that was published in a journal Oncogene. So it was right when Praveen had reported, we reported, that histone acetylation is a hallmark…so when you have HDAC inhibition, whether it’s the clinically used drugs that are used now in patients, things like vorinostat or these dietary factors, if they’re HDAC inhibitors, the first thing you need to do in order to show evidence of that is let’s look at histone and the histones should become acetylated in response to this treatment. And sure enough, we showed that numerous times in different types of cancer cells. What was interesting is that when Praveen did a time course, sure enough at 12 and 24 hours and later we saw histone acetylation.
But early on, the surprise was when we took all of the cells early on at 6 hours, colon cancer cells, and then did a proteomics approach—unbiased screening of all the proteins—before any change in histone acetylation this protein CCAR2 was heavily acetylated. So it said to us, although histone acetylation may be important later on, this protein seems to be a very important regulator of something in this cell. And as Praveen has mentioned, that was right around this time, this paper came out in Oncogene saying that it’s an important oncogenic factor. And they showed beautifully that it was actually acting as what’s called a co-activator of Wnt signaling. There’s a protein called beta catenin, which is considered an important oncogenic driver for colorectal cancer. And when this protein CCAR2 associates with beta catenin, it makes that protein far more active in terms of what it’s doing as an oncogene driving genes like MYC and other oncogenic genes.
So the fact that what we showed actually is through the acetylation of CCAR2, through this treatment with the broccoli compound, that acetylation prevented CCAR2 from interacting with beta catenin. So this important co-activator even though it’s still there, it can’t function like it’s there, it can’t actually physically interact with beta catenin. So therefore beta catenin/Wnt signaling was significantly less active.
The HDAC inhibitor field has been around some time now, 15 years at least. So those types of drugs have entered clinical trials and they are showing some promise. So early on they look very good for patients with advanced cutaneous T-cell lymphoma. Those patients responded extremely well to this drug vorinostat and so it was quickly moved into other trials and other HDAC inhibitors have come on the scene. The issue with them is that they have some toxicity: some cardiotoxicity, [some other heart defects], myelosuppression is one of the issues. So the thinking is, one of the reasons is that currently these drugs are called pan HDAC inhibitors. So they’re in multiple HDACs in our body, [in our cells]. So they seem to be targeting all of them at once.
Christina Sumners: So you can get all these side effects.
Dr. Rod Dashwood: So you can have side effects. And one of the interesting observations about some of the dietary compounds like sulforaphane, they are not what we would call pan HDAC inhibitors. They don’t simultaneously hit every single HDAC. In the case of sulforaphane in colon cancer cells, the first preferred target, it seems to be called HDAC3, which is one of the important HDACs [that’s overexpressed] and seems to be playing a role in colorectal cancer development. So that part is being developed quite well.
As this is happening, as drug companies are trying to develop HDAC specific drugs to avoid some of the toxicity issues, this whole other class of agents was being developed called ‘Reader’ inhibitors. JQ1 is not an HDAC inhibitor, it’s called an acetyl ‘reader’ inhibitor. In epigenetics we talk about readers, writers and erasers, right? So you’ve got the histones, you’ve got those little tail sticking out. ‘Writers’ would be things like acetyl transferases that put an acetyl group on. ‘Erasers’ would be HDACs that remove them, and then the ‘Readers’ don’t do those things…they actually just go on and find the acetyl groups, dock onto them, and bring in other proteins to affect chromatin access and gene expression.
[Jay] Bradner and his group sort of developed the ‘poster child’ for this new group of inhibitors, called JQ1, and they designed that specifically to target a subclass of these acetyl ‘reader’ proteins called the BET family. And so although some of them have moved into clinical trials and again have shown some promise, again [dealing with] the issue of toxicity and how do we refine these, now second and third generation acetyl ‘reader’ inhibitors are being developed.
I think it’s too soon to say that we want to take a broccoli compound and mix it with JQ1 and give it to patients, at this point. We still need to know more about the risk benefit of that type of question.
Christina Sumners: You’re talking about risks, just eating the foods then might be the best approach because eating broccoli sprouts, eating broccoli is pretty safe. Not going to have too many side effects.
Dr. Rod Dashwood: So Dr. Talalay, who is a professor at Johns Hopkins, he discovered the compound sulforaphane many years ago and he did a lot of the initial work. He basically screened every fruit and vegetable he could think of. And he isolated this compound sulforaphane as what he described as a phase two inducing agent. This is a pathway in our body that’s responsible for detoxifying and eliminating toxins from our body.
So if you can induce that pathway, that’s “a good thing to do,” right? So out of that, he discovered sulforaphane. But he did some very elegant additional work showing, first of all, if you compare the kind of mature broccoli that most people think of when they go to the salad restaurant. So in that case, the frozen broccoli had less activity than fresh broccoli. That was the first observation and there was variation among different places that he purchased this. The real interesting observation though was the broccoli sprouts. So if you take a broccoli seed and you make the little sprout at three days, that is actually loaded with the precursor of sulforaphane. So actually sulforaphane is not present in the foods as sulforaphane, it’s present as a precursor. It’s called glucoraphanin. And so when you take a fresh, like broccoli head and munch on it, you’re opening up the cells in the broccoli head.
There’s an enzyme called myrosinase. It’s released and activated. So it ‘breaks off’ sulforaphane and makes it active.
Christina Sumners: Okay.
Dr. Rod Dashwood: And so he showed that the broccoli sprouts have a highly concentrated form of glucoraphanin. So basically what happens is that amount of glucoraphanin that’s in the seed stays the same amount, but as the seed becomes the broccoli head, it’s diluted by biomass. So you could take one very large bowl of broccoli heads, or you could eat a half a cup of broccoli sprouts and you get the same amount of glucoraphanin precursor.
Tim Schnettler: See, and I was going to ask, would cooking it affect it? Did it, were there any studies on that, on cooking the vegetable versus raw?
Dr. Rod Dashwood: So that enzyme can be inactivated by cooking. And so, you know, if you eat raw, like some people like to munch on raw broccoli?
Tim Schnettler: Right.
Dr. Rod Dashwood: So that will activate it. So depending on where you eat and how you cook, it can be released in the stomach, in the small intestine or in the [lower] gut. And there are some gut bacteria that are thought to actually help release some of the sulforaphane as an active compound. The one observation I’ll make is, I mentioned, I’ve been in Japan on sabbatical. So over there if you have sushi at a high-class restaurant, instead of giving you this wasabi paste from a plastic tube, they would actually give you a wasabi root, and the little personal grater.
Tim Schnettler: Okay.
Dr. Rod Dashwood: Then what happens is when you grate that, you’re breaking open the leaf [root] and activating myrosinase. So that’s like mega-active compound, that’s why it’s so active when you eat it in that situation.
Christina Sumners: Because it is freshly ground.
Dr. Rod Dashwood: Freshly ground, yeah. Which is why if you prepare mustard from the powder, like yellow mustard, and eat it at that point on a hot dog, it’s very potent. You come back the next day, it’s not, right? Because a lot of the active materials has come off.
Christina Sumners: Interesting. So now you not only have to only think about what types of foods to eat, but how you cook them, how you prepare them, how long they’ve been sitting out.
Dr. Rod Dashwood: Definitely. Definitely.
Dr. Praveen Rajendran: So I just wanted to add something here in terms of the different ways people have been cooking broccoli. So there are studies that indicate that mild steaming of broccoli is not too bad in terms of the amount of sulforaphane it has. In fact, some of the studies show that mild steaming is actually even better. So somebody wants to make a broccoli rice, a broccoli carrot rice, and just mild steam it. It’s still retained. I mean it’s always good to take the sprouts and fresh plant. But mild steaming does best retain the amount of sulforaphane in the plant.
Dr. Rod Dashwood: Well I think it’s worth mentioning that there are a number of clinical trials around the country, and elsewhere, that are looking at this whole question. And I believe none of them to this point have actually used purified sulforaphane as the test compound in humans. In many trials, including the ones that we’ve done, we’ve taken the broccoli sprout extract, which we have validated, has a high level of sulforaphane content because of the way it was prepared. And then that’s made into a little pill and given as a supplement. Others have made it into a soup.
So for example, Tom Kensler who again, he was at Johns Hopkins working with Dr. Talalay many years ago, is now at the Fred Hutchinson Cancer [Research] Center. He’s done a number of very elegant studies in various parts of China looking, for example at liver cancer risk and sort of interventions that may be able to prevent this high-risk population from developing liver cancer. He’s also looked at lung risk, lung disease risk.
For example, in Beijing you have a lot of atmospheric pollutants and carcinogens, et cetera. So he’s done some very elegant studies looking at feeding just a bowl of broccoli soup, basically, standardized and amazing activity from the population over there saying, “Yes, we really want to be part of this trial, very motivated to be involved”. So it’s a great population to study, and he has some very interesting data from consuming a high broccoli soup with these sorts of compounds in them. It seems to be having beneficial effects in the circulation and elsewhere.
Christina Sumners: So is this an example of precision nutrition?
Dr. Praveen Rajendran: I think the word “precision” is now coming in vogue in terms of precision medicine. So that is basically looking at the molecular entities that are responsible for a certain effect and trying to extract those active ingredients and make them more specific to the patient population. So you’re trying to target a certain patient population and then you have a certain target in mind and you have a compound that can go and target the particular protein, for example here. So that concept is generally being referred to as precision medicine in terms of drug development. They’re trying to make, for example, antibodies that are very specific for protein and in wanting that patient population, that over-expresses the protein, right, so you call this as precision medicine. So you precisely trying to target that target population. I believe you can do the same thing with nutrition as well because the targets, as long as we can identify what targets are important here…
In this particular case we identified CCAR2 as an important target that is very high in colorectal cancer patients and we know that compounds like sulforaphane can now target CCAR2. But I think the very interesting part of our study is really the epigenetic combination part, which is what really gets me excited about. So for some reason we really don’t know exactly how, but when you combine these two different epigenetic compounds, sulforaphane from the diet and JQ1, which is a chemical, there is a lot of synergy in the way they work together. And so at least in the patient population that over-expresses CCAR2, we can clearly use this combination in clinical trials and there should be some synergy in terms of response. So yes, definitely, precision nutrition is something that is going to be more and more employed I think. What do you think, Rod?
Dr. Rod Dashwood: I think that’s a really good explanation. Just to follow up, again precision medicine is something we see… We have collaborators at MD Anderson Cancer Center, in the Department of Clinical Cancer Prevention. So we get a lot of our clinical samples from collaborator, Dr. Eduardo Villa Sanchez. And so one of his jobs, he sees patients with colorectal cancer, especially high-risk cases, they’re called FAP and Lynch syndrome. So one of the things you do in patients who present with a colon polyp is you do a genetic analysis. So depending on a panel of genetic changes that may or may not be present in that particular case, you would decide, okay, this drug is appropriate to give to this patient because they have that mutation rather than this other mutation. And so again, the same concept is applying for nutritional and botanical and natural samples. At least it’s still again in the early development phase, but it’s more towards sort of patient-oriented targeting of what you’re trying to do.
And again, Praveen mentioned that in the specific case of CCAR2, there is clear evidence that patients who have higher expression of CCAR2 in their colon cancers have worse prognosis. So their overall survival is much worse. So if you were to, say, take a hundred patients with colorectal polyps and screen them and the 30 of those patients have high CCAR2, those might be the ones that you would want to think about giving this type of combination epigenetic treatment specifically going after CCAR2, whereas the ones with low CCAR2 maybe something else would be better.
Christina Sumners: Would be a better treatment for them. Okay.
Dr. Rod Dashwood: Right. So it’s trying to use the genetic and epigenetic changes that you’re seeing in the tumors to try and target your treatment or prevention approaches more precisely. It’s still very much in the early days, but I think that’s the ultimate goal here.
Dr. Praveen Rajendran: And just to add onto that, Christina.
Christina Sumners: Yeah.
Dr. Praveen Rajendran: So in a disease like cancer, I mean it’s not one disease, a lot of people think that it is one disease. I mean the cancer has more than a hundred different types of cancers and then each patient’s cancer is his or her own cancer. So each cancer is different from what the other person has. Which raises a very important question. Do you have to treat the patient here like an individualized therapy?
So that’s why I think more and more of the clinical trials in cancer are focused more towards the patient. What characteristics are different, because none of us look alike. So each patient’s disease, is also very unique to that particular patient. So I think precision medicine and precision nutrition are going to be the treatment options for the future.
Dr. Rod Dashwood: And I think one other part of that, Praveen, is that we also know that even within one patient with one colon polyp, we have what’s called tumor heterogeneity. So that’s the concept that has really evolved over the last few years. So now you look at a particular cancer, it’s not like one or two or even five genetic changes. If you take different parts of that same tumor, you can identify different genetic changes and different epigenetic changes. So now how do you decide what particular therapy or therapies to do? [I think] this is why a lot of people are focusing more and more on the prevention side of things. If you can act earlier on where only one or two of these things are dysregulated rather than 10 or 20, you might have a better shot at preventing early stages moving to late stages, and maybe even reversing them before they go further.
Christina Sumners: So just going back to prevention. So that’s kind of really where we need to be thinking is how to prevent these types of cancers from forming. How do you see compounds in foods like broccoli, broccoli sprouts as helping with this?
Dr. Rod Dashwood: So this is something that Dr. Michael Sporn has had to address many years. He’s the person who first coined the term “chemoprevention”. And for many years it really was a very heavy focus on different dietary factors, phytochemicals, micronutrients, vitamins, things like that. And they’ve been studied every which way you can study them.
And so it depends on which side of the coin you fall. So people like me who are an optimist say that these dietary compounds are acting on different stages of the carcinogenic process. And so that’s a good thing. They’re effective because they’re not only targeting one dysregulated pathway, but potentially can target others as well. So multiple targeting of cancer steps. The flip side to that, however, is, well, if having these multiple steps, aren’t there also ‘off target’ effects? If that other pathway that they’re targeting isn’t altered or dysregulated in a particular cancer, isn’t that going to be an issue?
And so the term that’s often used is ‘pleiotropic’—dietary factors are pleiotropic; they have lots of different effects. Dr. Sporn has argued that if you use the same kind of arguments for cancer therapy drugs, none of them would be developed for the trials and none of them would have been used. But of course the argument there is that you’ve got a preexisting condition, a preexisting disease, you have to do something about it. You’re not going to use that type of chemotherapy drug as a preventive agent, obviously.
Christina Sumners: Right.
Dr. Rod Dashwood: So how can we think about dietary factors? I think it’s still, even after so many years of research, something we are trying to pursue and it’s all about dose-response and understanding mechanisms. If you don’t know what the mechanism or mechanisms are.
And I would argue that I think one of the issues that has sort of bogged the field down in a sense is people focus on their one ‘pet’ mechanism. Their one pet pathway/signaling mechanism – they find something going on there and “that’s all” they need to do. That’s it though…we’ve shown it inhibits that oncogenic signaling pathway. That’s it. And the question I think that needs to be asked more frequently is, well what about other things other than that pathway that’s your favorite Wnt signaling pathway? We need to understand these other potential pros and cons of these dietary factors. If we can do that, understand more about the complex mechanisms, maybe we can apply them in a better way for prevention and treatment.
Christina Sumners: Well, that would be exciting if everybody had a personalized list of the foods they need to be eating to prevent cancer. That would be amazing. Well, thank you both so much for joining us today. I don’t know about you, Tim, but I’ve learned a lot.
Tim Schnettler: I did. It was very interesting.
Dr. Praveen Rajendran: It’s my pleasure. Thank you.
Tim Schnettler: Appreciate it.
Dr. Rod Dashwood: Thank you.
Christina Sumners: And thank you all so much for listening, and we will see you next time.