It’s been a while since I talked about RNA interference here. It’s still one of those tremendously promising therapeutic ideas, and it’s still having a tremendously hard time proving itself. Small RNA molecules can do all sorts of interesting and surprising things inside cells, but the trick is getting them there. Living systems are not inclined to let a lot of little nucleic acid sequences run around unmolested through the bloodstream.

The RNA folks can at least build on the experience (long, difficult, expensive) of the antisense DNA people, who have been trying to dose their compounds for years now and have tried out all sorts of ingenious schemes. But even if all these micro-RNAs could be dosed, would we still know what they’re going to do?

A report in the latest Nature suggests that the answer is “not at all”. This large multi-university group was looking at macular degeneration, a natural target for this sort of technology. It’s a serious disease, and it occurs in a privileged compartment of the body, the inside of the eye. You can inject your new therapy directly in there, for example (I know, it gives me the shivers, too, but it sure beats going blind). That bypasses the gut, the liver, and the bloodstream, and that humoral fluid of the eye is comparatively free of hostile enzymes. (It’s no coincidence that the antisense and aptamer people have gone after this and other eye diseases as well).

Angiogenesis is a common molecular target for macular regeneration, since uncontrolled formation of new capillaries is a proximate cause of blindness in such conditions. (That target has the added benefit of giving your therapy a possible entry into the oncology world, should you figure out how to get it to work well here). VEGF is the prototype angiogenesis target, so you’d figure that RNA interference targeting VEGF production or signaling would work as well as anything could, as a first guess.

And so it does, as this team found out. But here comes the surprise: when the researchers checked their control group, using a similar RNA that should have been ineffective, they found that it was working just fine, too – just as well as the VEGF-targeted ones, actually. Baffled, they went on to try a host of other RNAs. Reading the paper, you can just see the disbelief mounting as they tried various sequences against other angiogenic targets (success!), nonangiogenic proteins (success!?), proangiogenic ones that should make the disease worse (success??), genes for proteins that aren’t even expressed in the eye (success!), sequences against RNAs from plants and microbes that don’t even exist in humans at all (oh God, success again), totally random RNAs (success, damnit), and RNAs that shouldn’t be able to silence anything because they’ve got completely the wrong sort of sequence (oh the hell with it, success). Some of these even worked when injected i.p., into the gut cavity, instead of into the eye at all, suggesting that this was a general mechanism that had nothing to do with the retina.

As it turns out, these things are acting through hitting a cell surface receptor, TLR3. And all you need, apparently, is a stretch of RNA that’s at least 21 units long. Doesn’t seem to matter much what the sequence is – thus all that darn success with whatever they tried. Downstream of TLR3 come induction of gamma-interferon and IL-12, and those are what are doing the job of shutting down angiogenesis. (Off-target effects involving these have been noted before with siRNA, but now I think we’re finally figuring out why).

What does this all mean? Good news and bad news. The companies that are already dosing RNAi therapies for macular degeneration have just discovered that there's an awful lot that they don't know about what they're doing, for one thing. On the flip side, there are a lot of human cell types with TLR3 receptors on them, and a lot of angiogenic disorders that could potentially be treated, at least partially, by targeting them in this manner. That’s some good news. The bad news is that most of these receptors are present in more demanding environments than the inside of the eye, so the whole problem of turning siRNAs into drugs still looms large.

And the other bad news is that if you do figure out a way to dose these things, you may well set off TLR3 effects whether you want them or not. Immune system effects on the vasculature are not the answer to everything, but that may be one of the answers you always get. And this sort of thing makes you wonder what other surprising things systemic RNA therapies might set off. We will, in due course, no doubt find out. More here from John Timmer at Nobel Intent, who correctly tags this as a perfect example of why you want to run a lot of good control experiments. . .