At A Glance
Name: Tariq Rana
Position: Professor, biochemistry and molecular pharmacology, University of Massachusetts Medical School
Background: Professor, pharmacology, Robert Wood Johnson Medical School — 1993-2001; Post doc, University of California, Berkeley — 1991-1993; PhD, chemistry, University of California, Davis — 1990
An article in Nature and a presentation by RNAi pioneer Andy Fire is all it took to convince Tariq Rana that the gene-silencing technology would be the next big thing. Five years later, he is still working on RNAi and recently accepted a position on the scientific advisory board of RNAi-based therapeutics developer CytRx.
Rana recently spoke with RNAi News about his work.
How did you get involved in RNAi?
I got involved in RNAi when I read the first paper that came out by Fire and Mello, published in Nature, showing RNAi. At that time, I was back in New Jersey [at Robert Wood Johnson] and I asked [the seminar committee to invite] Andy Fire to come give a seminar. I was there when the whole thing started because after the seminar we were so excited and I was talking to my fellow colleagues and I told them how big of a discovery this is.
At that time, people didn’t know that yet — this was in 1999. I went so boldly, I said that after Watson and Crick, I think this is the biggest discovery in cell biology. People started laughing, saying: “If you really think that, then why don’t you work on it?” I told them: “Yes, I will.”
Why I got involved — one was Andy Fire’s visit. The second is that [when] I saw the [Nature] paper, my lab was working on … TAR RNA. TAR RNA structure is involved in the life cycle of the virus that causes AIDS, HIV-1.
I have a long history of studying RNA structure and quickly I figured: This RNAi, there must be something weird about RNA structure, because with DNA this doesn’t work. My interest in the phenomenon was that there is something very special about RNA structure, which is leading to [RNAi]. That’s why I got involved.
During the first two years, what happened is what I always tell students not to do — I ended up not following my own advice: When a kid is born, first they learn to crawl, then stand, then walk, then run. [The advice is] never take your science and go right for the run.
What I did was, since I was also working on the lifecycle of HIV, I was very excited to basically knock down this virus. But I made 300 nucleotide-long RNAs after this paper came out in 1999, and started to introduce them into human cells and cell lines. As we know, longer sequences will kill the cell. [At that time], we thought: Great. We can solve this problem if we can make PKR-minus cell lines. … That’s where we ended up losing a year or two.
Then, suddenly, we saw the two papers come out from Phil Sharp’s lab, showing that when you take fly extracts, the shorter RNAs are coming out 21 to 24 nucleotides. Then, a few months later, Tom Tuschl published his paper in Nature [showing] that 21-mer [nucleotides] can silence.
Still, after that, our interest continued and we ended up finding out which ends of siRNA you can cap, and how, and which end is important. We published that in the fall of 2002. … [In] our first paper … we showed what basic structural and functional features of RNA are needed to cause RNAi in human cells. … After that, you can see the kits coming out — RNA labeled with dyes, and so on and so forth.
That’s the history.
Where does that bring us to now? What sorts of RNAi-related projects do you have going on it your lab?
Right now, we are working on several issues. But just to go back — one fundamental question in our lab was, [given] that in the case of worms, the RNAi effect can continue and go on, what is the half life of RNAi in human cells? Do we have this enzyme called RdRP, which can activate [RNAi] and continue it?
We showed that [the answer is] no — you can cap the 3’ ends and RNAi can still work in human cells, showing that we don’t need RdRP activity for this one. For RdRP activity, you need 3’ [overhangs] to be free so that [the effect] can continue.
After that, we measured the half life of RNA. It turns out it’s [about] 64 hours. Then we went ahead and decided to continue, to modify RNA backbones and different side-chains, to understand the rules of what kind of functional groups are needed to cause RNAi in our cells.
What we were really shocked to learn is that in one strand of antisense, the guide strand, we changed everything from the 2’ [overhang] — the ribosugar to fluoroamino — and basically took away the [overhang] group. This strand still entered the pathway.
Based on that, we were very excited that now we can modify this RNA, where this won’t even look like RNA, and then still this can enter the [RNAi] pathway. What we are working on now is how we can enhance the lifetime of this.
We have now made some RNAs that can last in cells two to three weeks, to knock down the gene of our interest.
The second issue we’re are working on — and I think papers will be coming out soon on this — is that we now wanted to link these RNAs with some small molecules or some other sequences. We don’t have to use the typical matter [that] people introduce into cells. If we have to come up with RNA and we want to inject it in mice, we can’t add all these compounds that people sell.
Now, we have RNA where we are labeling the different moieties and different small molecules to basically enhance the specific cell-type uptake issues.
The third project in the lab is that we’re trying to understand how this pathway is functioning in our cell system. We’re trying to use different structures of RNA, again, and different ways to understand which RNA will enter in the pathway and at how high an efficiency.
The fourth major issue in the lab is that if [there is] this machinery, RISC, which is there in human cells, what is the rule of this? And, in a given condition, how much RISC is there to continue the function of RNAi?
As you continue with your work, what are some of the challenges facing you?
The challenges are, as you have probably heard, getting this RNA into the cells when you deal with a live animal. So far, we don’t have a single matter in the field where we could IV a mouse and then see RNAi taken up by cells.
I think that’s the biggest challenge for the field.
This is something you’re trying to overcome yourself, or are you collaborating with someone?
We are working ourselves on this issue. We have created many siRNAs where we have them conjugated to different moieties — different small molecules and different sequences of peptide-like structures.
We have seen some uptake, but I feel like the next few years are going to be challenging.
Are there things that you’re not working on, but would like to get involved in?
Oh, yes. One question I would love to work on is this pathway of microRNAs. The interest will be in human ES cells, for example — embryonic stem cells — meaning what is the pool of microRNA that really controls the fate of these ES cell types.
That’s a question I am really fascinated about and would like to work on.
You only have so much time, though.
Exactly. We’ve got four or five different things … and there are only ten or 12 people.
You just joined the scientific advisory board of CytRx. How did you get hooked up with them?
The founder of CytRx’s subsidiary — it’s called Araios, although they changed their name to CytRx Labs — Michael Czech, I work with him on this campus. He told me what they are about to do and what their targets are, their vision, [and] where they are going, and I thought it was a perfect match.
Are you collaborating with any other RNAi companies?
No, not at the moment. I guess now I’m tied with CytRx.
Does the relationship with CytRx extend beyond being a scientific advisor? Is there a more formal collaboration?
No. This is just being a scientific advisor, at this point.
I guess I can see in the future, as this effort linking molecules to some moieties works out, I think they will be the first ones to be excited.