Thursday, December 13, 2012

Arrowhead and Alnylam Vying for Subcutaneous RNAi Delivery Success

The use of the intravenous route of administration for the currently leading systemic RNAi delivery technology, Tekmira’s SNALP technology, has been noted to be a drawback of the technology, especially for non-severe diseases and in therapeutic areas historically dominated by oral medicines (e.g. the cholesterol-lowering market).  As a result, the arrival of two delivery approaches that promise to allow for subcutaneous administration has been welcomed: Arrowhead’s Dynamic Polyconjugates (DPCs) and Alnylam GalNAc-siRNA conjugates which have shown data suggesting their clinical use for gene knockdown in the liver (at least initially; DPC with potential to go beyond the liver).  

A day ahead of Alnylam’s Roundtable on conjugate delivery, I thought it would be a good time to get into the mood and compare the two competing technologies.

Basic Chemistries

GalNAc-siRNAs consist of siRNAs to which a cluster of three N-acetylgalactosamine residues have been appended.  It is these GalNAcs that are recognized by the ASGPR receptor protein that is abundantly presented on hepatocytes.  The choice of three over just one or two GalNAcs is due to the synergistic binding of multiple GalNAcs to the receptor.

DPCs also comprise of siRNA conjugates, but involve an additional endosomolytic agent to facilitate siRNA release from the endosomes.  The two components can be mixed together so that the drug can be given as a single formulation.  This, however, also requires that both siRNA and endosomolytic agent end up in the same place.  For hepatocytes, this is achieved by conjugating the siRNA to a cholesterol moiety and the endosomolytic agent to GalNAc.  

The reason why two different targeting agents are employed are two-fold: reduced competition for the uptake receptor, and not requiring triantennal GalNAcs such as in Alnylam's case which seems to involve a quite costly chemistry. The reason why GalNAcs on the endosomolytic agent in DPCs are not so expensive is because as a polymer (a peptide in the latest versions) multiple mono-GalNAcs can be conjugated distributively and still achieve the same synergistic binding effect.

Potency and Safety

The Holy Grail in RNAi subcutaneous delivery appears to be to get formulations potent enough so that the desired level of knockdown can be achieved with volumes of 1ml or less: you can squeeze only that much liquid under your skin through a thin needle.

The first of Alnylam’s GalNAcs, ALN-TTRsc, achieves a 80% target gene knockdown (ED80) following repeat administration in preclinical animal studies.  This is below the (based on OTS 2012) 3mg/kg barrier that apparently would allow for 1ml or less volumes in humans.  What surprised me to see at the OTS meeting in late October was that the GalNAc potencies, both in rodents and non-human primates, varied quite a bit between the programs.  The TTR formulation actually had the poorest potency among the programs.  This surprised me even more so given that ALN-TTR01 and ALN-TTR02 (both SNALP programs) contained highly potent RNAi triggers.  In the case of PCSK9, ED50 of less than 0.1mg/kg were obtained.

It is possible that the differences are not just due to the natural sequence-specific differences in RNAi potency, but a result of advances in chemistry.  In particular, optimizing siRNA-conjugates for tissue/endosomal stability rather than serum stability as is often practiced in RNAi Therapeutics is critical.  Importantly, this consideration also applies to DPC technology.      

DPCs should be more potent than isolated GalNAc siRNAs.  This is because you are adding an endosomal release agent to the liver-targeted siRNA (e.g. GalNAc-siRNA) conjugate and unfacilitated release of nucleic acid out of endosomes is believed to be highly inefficient.

Arrowhead has reported various impressive potencies such as 99% knockdowns at sub-1mg/kg siRNA doses.  This to me is strong evidence of the superior potency of DPCs over GalNAc-siRNAs.  Moreover, it seems that DPCs may inherently require less frequent dosing compared to GalNAc-siRNAs for which Alnylam aims at weekly or twice monthly dosing.

What is unclear, however, is the amount and resulting safety and volume implications of the endosomal release agent.  In particular, the most impressive knockdown data seemed to involve saturating amounts of endosomal release agent (~6mg/kg).  The first-generation endosomal release agent, PBAVE, suffered from relatively high toxicity, partly as a result of premature unmasking in the blood.  It makes sense that the newer, ‘more natural’ peptide-based endosomolytic release agents are safer.  By contrast, assuming that the GalNAc sugar itself is harmless, I am not too concerned about the safety of Alnylam’s GalNAc conjugates.

In terms of potency, advantage Arrowhead, in terms of safety, advantage Alnylam.

Strategic Considerations

The challenge for Arrowhead will be to make the case of the benefit of increased complexity over GalNAc-siRNAs. Would the prospect of a 3- or 5-fold increase in potency e.g. be enough justification for the investment?  I say ‘prospect’ because Alnylam could obtain knockdown proof-of-concept data at least a year before Arrowhead, especially since Arrowhead is planning to conduct the first study with DPC (ARC-520 for HepB) in healthy volunteers and thus won’t be able to measure viral target knockdown.

In addition to potency, DPC has the important advantage that it may be a more widely applicable RNAi delivery platform.  This alone may tempt others to put some money down on the technology to see where it can go.

Although GalNAc-siRNAs and DPCs are currently clearly competing, there is also scope for them to synergize, especially in the area of oligonucleotide chemistry.  Curiously, Alnylam did seek access to DPCs earlier this year, supposedly for its evaluation in one of its 5x15TM programs.  Learning about DPC siRNA chemistry may be of at least equal, if not considerably more value to Alnylam.

Which of the two delivery technologies do you prefer for target gene knockdown in the liver?  Take the survey on the top right-hand corner.


kiesenbiker said...

Thanks Dirk for the blog. it's allways good to have a inside view from a specialist.

The press release from ARWR said:
Dr. Lewis’ presentation illustrates the efficacy of DPCs formulated for subcutaneous administration using Arrowhead’s latest proprietary polymer masking technology. Using DPCs to deliver siRNA, high-level target gene knockdown is observed at low siRNA doses without toxicity in rodents and non-human primates.

...doses WITHOUT toxicity ...

Is that not what it means to me ...WITHOUT is without?


RNAI inv said...

without toxicity

and you forgot the most important part

"in rodents and non-human primates"

Anonymous said...


Do you know what the volume of injection for ISIS mipo is for a dose of 200mg?

Dirk Haussecker said...

No drug without side-effects. If they have not seen serious side-effects so far, that's encouraging, I agree. But the human experience still has to be awaited.

Not sure about mipo dosing volume. My guess is that it is around 1ml though, or slightly higher (based on oligo solubility limits).

Anonymous said...

BG mentioned during roundtable that GalNAc is aimple one drug while DPC is more complex and involves two compounds. DPC may face greater FDA hurdel because of safety concern involving two compounds.

kiesenbiker said...

I do not think that ARWR with there DPCs will have a problem with the FDA because of there two compounts!
Even with four or five compounts its just the impact / output that counts. Its save or it is not save! It is allways the same procedure...
Science is defending the old standard because they are not able to imagine the new best in clas standards

Anonymous said...

Two components to the drug product will mean 2x as much of a headache for manufacturing and QC. But it's doable though.

kiesenbiker said...

From the ARWR Press Release:
This new delivery approach dramatically increases the efficacy of cholesterol-siRNA and, together with the co-injection strategy, simplifies the manufacturing process to enable a commercially scalable delivery vehicle for RNAi therapeutics...

So no headache for ARWR but for ALNY!

I understand BG (COO of ALNY)! I would say the same about the product of my competitor! To make ARWR product wicked, my product look better!

Anonymous said...

Dirk, your estimation "...Alnylam's case which seems to involve a quite costly chemistry. The reason why GalNAcs on the endosomolytic agent in DPCs are not so expensive is because as a polymer (a peptide in the latest versions) multiple mono-GalNAcs..." doesn't reflect the chemistry behind - this statement is wrong.
For the DPC system you have to synthesisze: (1)polymer (2)CDM-PEG (3)CDM-GalNAc (4) conjugate polymer/CDM-PEG/CDM-GalNaAc (5) RNA-Cholesterol
However, the synthesis of the tridentate GalNAc ligend and the coupling to RNA is straightforeward...just ask a chemist.

your statement "...Even with four or five compounts its just the impact / output that counts ..." doesn't take into account all the CMC issues that come along especially with such polymeric compounds, ever thought about that?... always underestimated.

Dirk Haussecker said...

Don't follow you really on the synthesis. I've heard from two independent sources that the tridentate GalNAcs are quite expensive to make. I also understand that DPCs do not require multidentate GalNAcs because you can get synergistic affinity from the various mono-GalNAcs on the polymer/peptide.

I also do not understand why there should be necessarily special CMC issues with polymers. The latest polymer in DPCs is a peptide. siRNA is a polymer/oligomer in that sense, too. Not a big deal IMO for siRNA and melittin peptide.

There are, of course, also more complex polymers such as fragmented heparins or the MS drug Copaxone by TEVA. But even here, as long as you can characterize a mix and manufacture that mix consistently, they can be turned into commercial drugs. Regarding DPCs, Arrowhead is apparently quite confident that it can overcome the manufacturing challenges now (esp. with the 2-component system) and get into the clinic. Having said that, Arrowhead in the past has not always met its guidance.

RNAI inv said...

Delivery is not that important, at least not yet as a platform. It's great that the tech gets better, but why stop here, next oral.

What the RNAI space really needs is products in phase 2/3 that will have benefits compared to other technology.

To me the LNPs and Galnacs have more value. Because LNPs are already in phase 2. And Galnacs because ALNY has the money to move this forward. Also in areas in which I think they have a good chance to get to the market.

So far it looks like big pharma is not really interested in preclinical delivery, especially without a target.

Anonymous said...

(1) ALNY's [GalNAc]3 is based on a synthesis already described by Sliedregt - most probably with some minor changes and improvements (Sliedregt et al. J. Med. Chem., 1999, 42 (4), pp 609–618.)The synthesis is straightforward: glycosylation is ß-selective, TRIS building block can be made in a few synthetic steps. Of course not trivial, but as you can see not a huge synthetic challenge.

(2)You are right, there are not necessarily special CMC issues with polymers. But keep in mind that once you have synthesized e.g. a melletin peptide (23-32mer), you conjugate i.e. steric stabilizer to the C- or N-terminal ends, furthermore you conjugate GalNAc ligands to polymer's primary amines - more than just one ligand, a couple of ligands and of course in a defined substitution pattern... But then you have to purify such polymeric conjugate and you have to characterize it - that is the most challenging job.

No doubt that ARWR can handle this and I do really like their approach - but we have to realize that between bench and bedside is a big hurdle called CMC, fortunately.

Anonymous said...

Is oral delivery really that important? Once you get to the potency when you need only once a month subc injection, then who really cares for oral delivery. As a patient I would rather take once a month subc injection and not worry about taking an oral pill everyday.

RNAI inv said...

Oral always is better than SC. If they ever make it work. And it certainly will not be a pill every day with oral.

On another note. ARWR will be running out of money again soon. More free warrants for anybody who gives them some money.

Anonymous said...

"more free warrants..." - could you explain this? Will they not simply issue more stock?

RNAI inv said...

So far they keep giving away free warrants with the new stock they issue.

They have trouble getting enough funding, so they have to give away something to get new money. This will limit any future potential for this stock, but short term it means the company doesn't have to do terrible discounts with the new stock.

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