RNA therapeutics: has the code been cracked?

RNA therapeutics have been around for a long time. Stanley Crooke founded Isis Pharmaceuticals (diplomatically renamed Ionis in 2015) in the late 1980s. They got an approval for treating CMV infections in AIDS patients in 1998 (a commercial flop)…and didn’t get a second approval until 2013. I don’t know the investment history of Isis, but they must have burnt through billions of dollars along the way.

Isis used antisense technology, which has a very simple mode of action: short RNA sequences bind to mRNA from specific genes. Cells don’t like double-stranded RNA (because they look like viral genomes) and cut up the mRNA, effectively silencing the target gene.

Once you have the gene sequence, all you need for an effective therapeutic is to get the antisense RNA into the cell. How hard could that be?

Plenty hard, as it turns out. Cells don’t allow many large molecules inside, especially molecules like nucleic acids–which may well be viral invaders. And even getting an RNA to the cell is incredibly difficult. Plasma is full of nucleases that cut up RNA, the liver sucks it up like a sponge, the kidneys filter it out almost immediately. The half-life of an RNA injected into the bloodstream is on the order of seconds. Most biologics (large therapeutic molecules like enzymes and antibodies) have half-lives of days or even weeks.

Other nucleic acid therapeutics faced the same problems. I was one of the first scientists hired at NeXagen (later NeXstar), founded in 1991. We used aptamer technology to create RNAs that bound to protein targets and neutralized them, much like antibodies do. Aptamers can work on extracellular targets, so we had one less hurdle than antisense. But the clearance problem was still a killer, particularly as we wanted to focus on oncology and inflammatory diseases–chronic diseases that required days to months of therapy.

One of my assignments was to solve the clearance problem. I came up with the idea of conjugating aptamers to albumin, an abundant and long-lived serum protein that carries many small molecule drugs. This, along with some modifications that reduced degradation, increased aptamer half-life from 30 seconds to about 30 minutes. But my bosses hated the conjugation idea and 30 minutes was still too short. I believe I got no bonus and a minimal raise that year and was passed over for promotion in consequence of my failure.

That was in 1994. As of 2018, nobody had done much better in solving the RNA clearance problem. RNA therapeutics, despite a couple of approvals, was still very much a fringe technology and a huge money-loser. Merck bought Sirna Therapeutics (right next door to NeXstar) for $1.1B in 2006, thinking the clearance problem was solvable. In 2014, they gave up and dumped it for about 15 cents on the dollar. Despite a couple of niche approvals here and there, mostly for liver diseases, RNA technologies have been most useful as basic science tools and as a proteomics platform.

That may soon change. After 20+ years of nobody solving the clearance-and-cellular-uptake problem, maybe somebody has. That somebody (or rather somebodies) are a group at Northeastern University and MIT.

Their modifications to a siRNA increase its plasma half-life from minutes to hours:

The modified RNAs are taken up efficiently by cells in culture, and by tumors in mice:

And the uptake of the cleavable form of the RNA-polymer conjugate (pacRNAClv) is sufficient to suppress tumor growth:

I don’t mean to imply that these results show that this siRNA formulation could be a cancer treatment breakthrough; lots of failed drugs show similar or better results in animal models. But for the first time, RNA therapeutics are in the game. They now have to be considered a serious therapeutic platform for cancer and other chronic diseases.

What is this miracle polymer you ask? It’s boring old PEG, polyethylene glycol. PEG has been used for decades to improve the pharmacokinetics of aptamers and proteins. We used it at NeXstar, and it helped quite a bit (although not nearly enough). PEG is normally synthesized and conjugated as a simple linear polymer:

The two-carbon one-oxygen motif is repeated hundreds of times to form a very soluble, flexible and non-toxic polymer that sucks up water and increases the size of whatever it is linked to. The innovation of Xueguang Lu and colleagues is to make this polymer highly branched, so that (as the article title says) it resembles a bottle brush. The resultant brush polymer more or less creates a cloud of water and PEG around the RNA, protecting it from nucleases and clearance organs like the liver and kidney, and promoting its uptake into cells. It looks like this in graphic simulation:

This is one of those innovations that seems obvious in retrospect. It was certainly doable 20 years ago, but nobody thought to try. The introduction section of the paper has a beautiful and concise summary of all the problems and failures of previous approaches, which alone makes the paper worth reading. More subtly, of course, the authors are taking a victory lap. You get to do that when you solve a decades-old problem.

It remains to be seen how general this approach is. Although the authors claim that the protected oligos are still able to hybridize with complementary strands of DNA, it seems that protection from nucleases implies the exclusion of other proteins as well, including therapeutic targets. That would be a problem for aptamer applications.

But even if bottle brush PEG is not the ultimate solution to the RNA PK problem, it’s a big advance in a field that hasn’t really seen one since…ever. It may very well open up the field of RNA therapeutics. Maybe we’ll even find some use for all that genome data we’ve churned out, data that has mostly been of no medical value. That would be quite an accomplishment.