Anti-miR122 Antagomir Successful in Fighting HCV in Chimpanzees

I just wanted to post this video by KENS 5 on the anti-miR122 studies in HCV-infected chimpanzees by Danish LNA company Santaris and researchers from the San Antonio Southwest Foundation for Biomedical Research. The study, just published online in Science, shows surprisingly potent reduction of HCV levels (2 1/2 logs) with an miR-122 antisense inhibitor and, at least equally important, without the emergence of viral escape mutants. While this study only involved four chimpanzees and human studies are still in the early phases (phase I, healthy volunteers), these results justify hopes that such a first-in-class agent could be a valuable component of future combination therapies with increased cure-rates, shortened treatment times, and maybe without the need for current standard-of-care component ribavirin.

Curiously in light of these promising results, it appears as if GSK let an option to the Santaris compound expire. While Big Pharma may have multiple reasons why it does not want to pursue a certain compound aside from the science (e.g. the fact that HDL-cholesterol was lowered as a result of miR-122 inhibition could warrant caution), for example product portfolio considerations, one possibility is that IP concerns and GSK's relationship with Regulus were important factors: Regulus has an exclusive license to the IP surrounding the fundamental work by Jopling and Sarnow on the role of miR-122 in HCV replication. While this may be good news for Regulus, I was made aware that the broad Esau and Tuschl patent applications on microRNAs as therapeutic targets and held by Regulus are being narrowed down considerably by the patent offices.

Note: For a more detailed discussion of this paper, please read the review by Mark Kay and myself in Molecular Therapy (click here).

RNAi Therapeutics 2010: The Year of the SNALP

While the broad markets have enjoyed a considerable rally since the lows earlier this year, with the S&P500 up more than 65% since March, shares in RNAi Therapeutics companies have only initially participated, but then reversed course. This is unfortunate since this does not make establishing a broad drug development platform any easier with no products on or close to the market. I want to be clear though that one cannot blame it all on a market that 'does not get it' or just bad luck: strategic mistakes have been made, false expectations raised to a point that the market, and this might include Big Pharma, is saying ‘show me credible non-human primate, or even better, human data before I believe you’.

On the other hand, chaos brings with it opportunities, especially for companies that can emerge from this confusion with pre-clinically well-validated technologies and unambiguous proof-of-concept data for therapeutically relevant gene knockdown in humans. A lot will therefore depend on whether a single delivery technology, SNALP, can achieve such results. Results from both SNALP-ApoB (Tekmira), expected at the end of Q1 2010, and ALN-TTR (Alnylam) later in the year, provide opportunities for demonstrating efficacy in relatively small patient populations. Safety, of course, will be equally important to watch.

There are other RNAi Therapeutics candidates in the clinic among which maybe Benitec’s HIV program may provide molecular indications of antiviral activity with the rHIV-shI-TAR-CCR5RZ triple RNA(i)Rx combo. Cancer-related clinical results will mostly focus on safety, although ALN-VSP02 results could go into more mechanistic depths. Quark Pharmaceuticals’ candidates, of course, are far ahead of the field- sometimes I ask whether possibly too far ahead in light of what we have learned about the uptake of naked siRNAs and innate immune stimulation. Beyond RNAi Therapeutics, progress with mipomersen, DMD exon-skipping, and miR-122 inhibition for the treatment of HCV could help return optimism to RNA therapeutics drug development in general.

Taken together, I believe that 2010 could indeed be remembered as the RNAi Therapeutics Year of the SNALP, although it is always possible that a MEGA-deal, possibly inspired by the ApoB-TTR results could divert some of the immediate attention. Some of you may remember that I called out 2008, also for reasons related to SNALP, as the RNAi Therapeutics Year of the Liver. I still believe that this would have been possible if the attention had been focused properly on the exciting development path of this technology for liver applications some of which are now entering the clinic, instead of the somewhat broader messages the market received and is now struggling to cope with. After SNALP, cancer is a strong runner-up, and may in fact drive some of the major business developments of 2010. 2011 or 2012 may be the RNAi Therapeutics Year of Cancer outright.

Given my obvious fondness for liposomal delivery and to stay on top of the exciting scientific developments in this area, e.g. targeted delivery, I am already looking very much forward to be attending the annual International Liposome Society meeting in London next week. All the while next door Cancer RNAi Therapeutics company Silence Therapeutics, which also works on somewhat related lipid-mediated siRNA delivery, should be discussing their merger at the General Meeting and is just one more reason to go.

RNAi Therapeutics Portfolio Update

As we approach the New Year, I decided to take a look at the RNAi Therapeutics portfolio and finally take out gene therapy company Oxford Biomedica. This is not because I have lost faith in gene therapeutics, recent clinical data strongly suggest otherwise and the ocular/neuro applications approach that Oxford Biomedica takes, also in partnership with Aventis, should make this one of the companies in the field to watch. Oxford Biomedica, however, has done too little in RNAi Therapeutics drug development to justify its place in the portfolio. I still wonder how ocular DNA-directed RNAi Therapeutics for example could be institutionalized- maybe as part of a more general gene therapy company such as Oxford Biomedica, or an eye-focussed RNAi Therapeutics company employing both synthetic and ddRNAi techniques. Maybe even packaged into a re-formulated Targeted Genetics, yet another company that provided clear gene therapy clinical efficacy data for a rare eye disease. Until more strategic clarity is provided, however, including their continued interest in RNAi Therapeutics or not, I decided to sell some of TGEN as well.

The proceeds from these sales were put into ISIS Pharmaceuticals whose shares I believe have been oversold in the wake of the somewhat lukewarm phase III homozygous FH mipomersen results. Considering previous clinical results obtained with mipo as well as the overall favorable lipid profile changes as a result of ApoB knockdown, chances are that the upcoming phase III results in the other severe hypercholesterolemic populations will look better and signal the start of the manic phase of the manic-depressive mipo story.

Otherwise the portfolio should be well exposed to the potentially major value creating force in RNAi Therapeutics 2010 with Tekmira now making up the largest position in the portfolio, largely the result of the relative weakness in ALNY (-40%). Silence Therapeutics should warrant a re-evaluation after they make public their merger partner. AVI Biopharma remains on the radar for their involvement splice modulation (DMD foremost) and other areas that could provide them with near-term revenue, especially if they should move closer with mdRNA which could make for an attractive combination. Just fresh from the press is also the announcement that ISIS Pharmaceuticals will play a more active role in splice modulation.

Has mdRNA Found a Way around Alnylam’s RNAi Trigger IP Wall?

mdRNA has to be The RNAi Therapeutics corporate success story of 2009. At the brink of bankruptcy at this time of last year after problems with their former nasal delivery business sent the company into a breathtaking tailspin, the new management, critically made up of former Sirna Therapeutics executives, was able to save the company by quickly cashing in on some non-core assets, attracting upfront payments of about $12M in two non-exclusive RNAi Therapeutics deals with RNAi heavy-weights Novartis and Roche, and an opportunistic financing shortly after final approval of a partnered legacy generic (calcitonin-salmon nasal spray for osteoporosis) caused an irrational spike in mdRNA’s share price. Although I perceive mdRNA’s lack of RNAi Therapeutics publications as a clear weakness, also in light of its history of later unsubstantiated scientific claims, these developments are signs that not only is the management apparently well connected, but that the company has been able to establish a quite decent RNAi Therapeutics drug development platform and with IP that cannot be dismissed out of hand. Since the Novartis deal on delivery involved not only IP, but also the transfer of liposomal formulation know-how, but the Roche deal on RNAi triggers appears to be an IP-only deal, I have looked more into mdRNA’s claim of having freedom-to operate with its ‘proprietary’ RNAi trigger designs. Out of fairness, I should add here that a lot of this change had been set in motion with the old management.

It has become a popular game in the RNAi Therapeutics industry to design ways around Alnylam’s dominant RNAi trigger patent portfolio. While corporate strategy is the driving force behind these supposedly proprietary designs, given that intellectual property is not always guided by the spirit of science and since new trigger designs might have unexpected beneficial properties for certain applications, it is prudent to pay attention when Roche is willing to pay mdRNA non-refundable $5.0M for a non-exclusive license to mdRNA’s RNAi trigger IP claims after having just paid more than $300M for much less of Alnylam’s RNAi trigger IP. Of course, one might also view this deal, which was also largely free of downstream obligations, as a move by Roche to cover all their bases- just in case.

At the time of that deal, mdRNA had three RNAi trigger designs in its stable: certain rights to City of Hope’s Dicer substrates, the three-stranded meroduplex siRNA design after which mdRNA takes its name, and unlocked nucleic-acid-modified siRNAs (usiRNAs). Subsequent to that deal it emerged from regulatory filings that, perhaps because Dicerna appears to own most of the Dicer-substrate IP, that mdRNA has dropped Dicer substrates from their portfolio. Furthermore, since Dicerna would probably have been quite happy to make a deal with Roche, it would appear that Dicer-substrates, although part of the deal, was not the main motivation for Roche. This leaves us with meroduplexes and usiRNAs.

If you assembled five PhDs in molecular biology for a weekend, presented them with the main claims of Tuschl I and II, and asked them to come up with RNAi trigger design-arounds, in principle the result may have looked similar to what mdRNA arrived at. That the various designs should also likely to be functional biologically, at least to a certain degree, is a testament to the robustness of the RNAi pathway when it comes to relatively short double-stranded RNAs, a realization that has only sunk in subsequent to the Tuschl publications, but that was already heralded by the findings of Kreutzer and Limmer.

Both Tuschl I and II essentially claim double-stranded RNAs of 21-23 (T-I) or 18-24 (T-II) (contiguous?) NUCLEOTIDES in length. Besides size and overhangs which had been the main theater of the RNAi trigger IP war thus far, one might argue that an RNAi trigger that does contains a number of NON-nucleotides or in which the dsRNA is made up of three, and not two strands, does not literally violate the Tuschl claims.

I have long been puzzled why mdRNA would think that using unlocked nucleic acids (UNA) per se would be the solution, as I considered UNA just another nucleotide-modification alternative in the armamentarium of the siRNA synthetic chemist: UNAs are simply nucleic acids in which the C2-C3 bond has been disrupted (that is apparently also the process by which they are generated). But when the company’s CEO Michael French at the BMO Capital Markets Focus on Healthcare Conference made a point that it is the fact that UNAs supposedly are not nucleotides is indeed what underlies mdRNA’s interest in UNAs. A quick, non-representative survey among scientists that I know, however, was consistent with my initial assumption that UNAs clearly have to be considered ‘nucleotides’ as they look and behave like such. Moreover, there are numerous quotes by the inventor of UNA, Jesper Wengel of Denmark, and by the Danish company RiboTask from which mdRNA has exclusively licensed the UNAs for therapeutic applications that UNAs are modified nucleotides. The question therefore appears to boil down to whether potential partners first and then the patent offices can be convinced of that a ‘nucleotide’ that does not contain an intact ribose or deoxyribose strictly is a nucleotide no more. Although I am skeptical that any one nucleotide modification is sufficient to get around the Tuschl patents, I could imagine that the possibility alone of their literal interpretation could motivate others to come up with similar non-nucleotide siRNAs.

Aside from the IP considerations of usiRNAs, it is premature to judge the scientific value of this modification over others. The fact that ‘unlocking’ a nucleotide is a quite distinct modification should allow it to endow an siRNA with unique properties with respect to recognition by the RNAi machinery, target specificity, the potential to be recognized by innate immune receptors etc. What can be said based on the limited publication record (Bramsen et al., 2009: Large-scale screen....; Kenski et al., 2009: Analysis of acyclic nucleoside modifications...), UNAs like many other modifications are best tolerated in the passenger strand and can reduce off-targeting by the passenger-strand, whereas the use in the guide strand is much more position-dependent. Whether its judicious incorporation can indeed largely abrogate innate immune responses and/or allow for a better differentiation of on-target cleavage over microRNA-like off-targeting as claimed, remains to be seen and especially published (also because mdRNA/Nastech has a track record of making similar claims which tend to silently disappear over time). On the other hand, since Sirna Therapeutics (Merck) is evaluating UNA-modified siRNAs (Kenski et al., 2009), despite its access to Tuschl I, may be interpreted as a sign that there may indeed some unique scientific merit to usiRNAs, and mdRNA may derive value from controlling its use for RNA therapeutic applications.

Moving on to 3-stranded siRNAs, the other pillar of mdRNA’s RNAi trigger strategy, from an IP point one has to probably say that this stands a better chance of surviving Tuschl scrutiny with Tuschl II explicitly claiming double-stranded RNAs consisting of two strands. The double-strandedness of Tuschl I, however, could be more broadly interpreted in that a nick should not make a difference in determining the length of a double-stranded region. In this case, it will be interesting to find out how 3-stranded siRNAs with a gap instead of a nick would perform.

When I first heard of the 3-stranded siRNAs in 2006, I thought that this was based on the observations by a number of labs at the time that the passenger strand of the siRNA is cleaved as part of RiSC activation and that the objective of 3-stranded siRNAs thus was to hi-jack the RNAi machinery downstream of the intact siRNA. However, looking at the history of the relevant patents, it appears that this was not the case, but was merely a coincidence. There is, however, an interesting twist to the story. RiboTask, which happens to also be mdRNA’s partner in usiRNas, filed for almost identical patent protection, but with a priority date that appears to be a couple of months ahead of mdRNA’s. In the case of RiboTask, which calls the 3-stranded siRNAs not meroduplexes but small internally segmented interfering RNAs/LNAs (sisiRNA/sisiLNA), I do not want to be as dismissive about their scientific motivation, as they address a major concern of 3-stranded siRNAs related to their potential instability. Since short segments of double-stranded RNAs should be relatively unstable, they demonstrated (Bramsen et al., 2009: Improved silencing properties using small internally segmented interfering RNAs) that it is possible to restore ‘duplex’ stability by modifying it with locked nucleic acids, the mirror-image of UNAs and curiously invented by the same people. Moreover, they show that while heavy modification of both strands renders most siRNAs inactive (with the exception of 2’F and 2’-O-methyl), converting such a modified siRNA into a 3-stranded siRNA often restores at least some of the knockdown activity (also quite interesting from an RNAi mechanism point of view). The ability to heavily modify an siRNA without losing activity may be particularly useful for applications where the siRNA is exposed to nucleolytic degradation such as for conjugate-siRNA approaches. From a scientific perspective, I should also mention that the 3-stranded siRNA design is a way to abolish passenger strand microRNA-type off-targeting, although this is no more a considerable challenge for RNAi Therapeutics. It should also be of interest to evaluate how nicked, and particularly gapped siRNAs are recognized by the various innate immune receptors. The above demonstrations may also help RiboTask and mdRNA in their way through the patent offices, particularly in the US and in light of the mentioned academic research that might have started before RiboTask’s work, and I am curious whether the usiRNA relationship between RiboTask and mdRNA will be extended also to sisiRNAs/meroduplex technology.

Since IP is subject to interpretation and therefore fraught with some uncertainty, it remains to be seen whether mdRNA can achieve certain freedom-to-operate in the RNAi trigger space. Although usiRNAs are the current focus of the company, keeping the meroduplex as an alternative design option open seems prudent. More broadly, RNAi Therapeutics in general can only gain from investments in evaluating new modifications and designs as unforeseen beneficial properties could emerge from this. Until then, however, it is unlikely that any Big Pharma would want to risk their RNAi Therapeutics future entirely on mdRNA’s RNAi triggers. This also explains why mdRNA sees more partnership potential in its RNAi delivery efforts, the currently more pressing need in RNAi Therapeutics and for which the financials should continue to grow. mdRNA urgently needs such a partnership since although the company had just averted bankruptcy, with cash running out over the next 3 months, the struggle to get out of the bankruptcy quicksand has not stopped. Based on management, while a number of early-stage technology evaluations are ongoing, no significant upfront payments from partnerships can be expected until the middle of next year by which time it hopes to have more non-human primate data to satisfy a more discriminating Big Pharma audience. Given the positive developments of the company over the last year and management’s track record, investor interest should be sufficient to support a PIPE within the next 30-45 days so that the company gets a shot at achieving its partnership goal. If one indeed wanted to make the gamble on mdRNA’s management, from an investment perspective, waiting until more is learnt about the terms of the financing may be prudent, unless valuations continue to fall much further.

Calando’s RONDEL RNAi Therapeutics Delivery Promising, but CALAA-01 Prematurely Entered into Clinic

[Part 1 of a 3-part collaborative series with Tobias Wolfram on the first notable attempts at RNAi Therapeutics for solid cancers that have entered the clinic]

When during the company's R&D day the CEO of Alnylam, John Maraganore, highlighted Calando’s cyclodextrin-based siRNA delivery technology (RONDEL) as one of the noteworthy non-SNALP systemic siRNA delivery technologies out there, it certainly piqued my interest. This is not least because any cash infusion and longer-term commitment by a partner like Alnylam would do wonders for the parent company of Calando, Arrowhead Research which is a conglomerate of early-stage, IP-focused business units and has just barely scraped by bankruptcy through a diet of cutbacks. While I always remembered the maturity of RONDEL, developed in the Mark Davis lab at Caltech, to be years behind SNALP, Tobias and I decided to to take a closer look at the development path of CALAA-01, the first clinical RONDEL delivery candidate and also investment focus of reorganized Arrowhead Research.

When Tobias first heard of the technology, it struck him as a very elegant, because simple, and modular method to formulate targeted nanoparticles. In fact, there are not many targeted nanoparticle siRNA delivery approaches where the components supposedly can be assembled by the pharmacist just before patient administration. RONDEL siRNA delivery consists of mixing together siRNA, a short cyclodextrin-containing polycation, and adamantane-coupled PEG stabilizers some of which carry a transferrin ligand, so as to create 60-80nm particles. These particles were rationally conceived to satisfy a range of pharmacologic and formulation considerations. Their suitability for solid cancer relies on the enhanced permeability and retention (EPR) effect of nanoparticles with reasonably long circulation times (here supposedly achieved by PEG stabilization), the ability of the particles to be taken up into cancer cells by transferrin receptor-mediated endocytosis and their subsequent release into the cytoplasm in a pH-dependent manner.

Unfortunately, what we soon came to realize was that while the concept is a very nice one indeed, the particles, particularly CALAA-01, remain to be better characterized both physically (shape, uniformity, storage and biological stability etc) and for their RNAi knockdown ability in vivo. For example, knockdown of RRM2, the target of CALAA-01, and subsequent tumor inhibition have not been demonstrated in a convincing in vivo system. Instead, knockdown efficiencies have largely been limited to in vitro studies. Moreover, these involved siRNAs that were selected with what today would be considered outdated methods and probably as a result were not very potent. The in vivo studies were essentially limited to pharmacological investigations, with the combination of in vitro efficacy and in vivo pharmacology forming the rationale for moving CALAA-01 into the clinic. Moreover, even when considering only the pharmacology, measures such as biodistributions and circulation times did not fit the model which may be explained by nanoparticle instability in vivo, something that really needs to be investigated further. Also, since the siRNAs were unmodified it strikes me as rather strange that no innate immune induction and only moderate adaptive immunity were reported.

What I found to be a valuable take-home message from those studies, although according to Tobias’ liking resting too much on indirect evidence, but not necessarily data obtained with the CALAA-01 clinical candidate, was that the utility of the targeting ligand appeared to be in increasing the cellular uptake of particles with little positive surface charge, less so in skewing the biodistribution towards the solid cancers per se. This could also be because the particles were cleared relatively rapidly from circulation, mostly into the kidney and bladder, which raises further questions about the purity and integrity of the particles. In this light, the mention of nanoparticle assembly by a pharmacist may also be interpreted a necessity due to storage problems of fully formulated particles. Nanoparticle assembly is notoriously sensitive to even slight changes in parameters such as temperature, speed of mixing etc so that it would be preferable for the physician to just administer the drug without the need for prior handling. It is therefore unfortunate that results from long-term storage and robustness of the formulation method were not presented. Nevertheless, an enhanced cellular uptake through the addition of a ligand could critically increase the therapeutic index of a cancer RNAi Therapeutic, and the modular nature of the RONDEL system should easily facilitate such additions.

Our assessment that CALAA-01 was probably entered into the clinic too early naturally rests on the publicly available data only. The publication dates of the relevant data, however, strongly suggest that they indeed represent the relevant data points and considering the financial situation of Calando, it would not appear that much were to be gained by holding back on positive data. This is by no means to belittle what otherwise is very rich science. Unfortunately, it is here that the tension between corporate demands for advancing a pipeline and the need to sufficiently advance the science is most evident and in the end risks harming both objectives. Optimistically, completion and evaluation of the CALAA-01 phase I trial will allow for valuable insights into the performance of RONDEL delivery in man for the benefit of any follow-up programs. Without any such strong data, it is questionable however whether the promise of RONDEL as a differentiated and flexible platform for RNAi Therapeutics delivery alone will be enough to make Arrowhead Research a good RNAi Therapeutics investment.

PS: To expand on the latter point, Tobias and I also discussed that, in general, it is easy to caution against entering RNAi Therapeutics candidates into the clinic early and dismiss such as a move of desperation. On the other hand, the case can be made that, when it comes to RNAi Therapeutics as a broadly applicable platform, clinically evaluating candidates which for example would not be expected to effect large knockdowns can be justified in that the data coming out of these studies may provide timely data invaluable for follow-up programs using very similar delivery approaches. Although investors will rightly fear the costs of a failed trial particularly for small biotech companies, such data may be valued more highly by a potential Big Pharma partner. This argument receives added weight in an environment like now where it is very difficult to raise capital from the public markets, and partnering is the primary means for small biotech to obtain capital at acceptable terms.

Merging Antisense with RNAi Therapeutics to Create Fitter Companies

Yesterday’s long awaited data presentation on phase III results from ISIS Pharmaceutical’s lead antisense gene knockdown program was met by disappointment in the investor community: ISIS down over 15% on the day. While the exciting news is that the drug, mipomersen, looks like it is very close for approval for the severe, but very rare condition of homozygous familial hypercholesterolemia (hoFH) and importantly also possibly other forms of highly elevated LDL-cholesterol, part of the disappointment may be related to the long-term outlook on antisense for gene/mRNA knockdown: a 27% reduction in ApoB levels following 6 months of 200mg weekly injections with a safety profile (a number of cases of elevated liver enzymes, injection side reactions) that may be adequate for the severe cases of hypercholesterolemia, but not necessarily for less severe diseases. And this is for an organ, the liver, which has one of the best pharmacologies for antisense. Also, while a 27% knockdown may be therapeutic for a few targets, especially in metabolic disease, for most other targets it is insufficient.

Lack of high-quality gene knockdown opportunities would lead to an inefficient use of its capital that includes an enviable $600M+ cash pile and an equally remarkable cash-flow from their ‘satellite businesses’. Maybe partly because of this realization, but also of course because it believes that its IP is transferable to all areas it considers antisense, ISIS is not standing still and continues to innovate in areas such as RNAi Therapeutics and the promising gain-of-function antisense technologies of splice modulation and microRNA inhibition.

On the other side of the fence, RNAi Therapeutics is also facing a challenging investment environment. Capital is particularly difficult to raise for early-stage platform technologies. While some companies such as Alnylam, Tekmira, Sirna Therapeutics (historical example), and mdRNA are making good strides in establishing broad RNAi drug development platforms, this alone does not make up for lack of steady newsflow from mid- to late-stage clinical results that can support rich valuations in biotech. This is also not helped by some obvious mis-steps within the industry itself with the main sins being spending money on lawsuits and prematurely entering programs into the clinic, ironically not least in an attempt to keep investor interest levels high.

By merging with antisense, both the pipeline maturity profile could be enhanced and resources spent more efficiently by avoiding the temptation of entering programs into the clinic prematurely and otherwise weeding out programs that serve to artificially fill pipelines. The latter point, of course, would also apply to the antisense company, and the antisense company would further benefit from gaining access to the most potent and therapeutically promising gene knockdown technology known, RNAi Therapeutics. As the intricate relationship between Alnylam and ISIS Pharmaceuticals or the acquisition of Coley as a launch pad for Pfizer’s RNAi Therapeutics ambitions demonstrate, the scientific barriers for such mergers should be relatively minor. One risk, however, that cannot be ignored, particularly for those companies that have been built for sale, is that by combining various drug development platforms, the new entity may become a less attractive candidate for a Big Pharma acquisition. When it comes to mere survival, however, a combination should be the lesser evil.

I will now briefly discuss three fantasy combinations, at least one of which I would speculate to see within the next year.

1) Alnylam Pharmaceuticals and ISIS Pharmaceuticals (probably not within the next year). Without a doubt the most influential and potent leaders in RNAi Therapeutics and antisense, respectively. Already highly entangled through their IP cross-licensing agreements and microRNA therapeutics spin-off Regulus, a combination would create an almost cash flow-positive dream team with over $1B in cash, blocking IP, unmatched expertise in nucleic acid chemistry, clinical pharmacology, and RNAi Therapeutics. First revenues from the sales of mipomersen and continued IP licensing revenues would support a solid pipeline consisting of mipomersen label extensions, RNAi Therapeutics opportunities for liver and solid cancers, full ownership of the miR-122 program for the treatment of HCV, and transitioning antisense towards splice modulation, microRNA inhibition (with Regulus), and possibly other gain-of-function antisense applications. As part of the re-organization, some programs could easily be sacrificed without punishment by the markets. John Maraganore would be the CEO of the combined company, allowing Stanley Crooke to follow his passion in the science of oligonucleotide therapeutics.

2) mdRNA and AVI Biopharma (how soon?). The transition of antisense for gene knockdown to gain-of-function applications is most noticeable in the case of AVI Biopharma. After many years of attempting gene knockdown with their steric-block morpholinos, the company finds success in applying its technology for the modulation of gene splicing. It is considered the closest competitor to Prosensa’s Duchenne Muscular Dystrophy exon skipping program which has only recently entered an attractive $25M upfront plus multimillion bio$$$ milestone and royalty deal with GSK after a series of high-profile publications on DMD-related exon skipping. The partnership potential of AVI’s DMD program then as well as its recent capital raising should provide the needed capital cushion for the combined company to invest in RNAi Therapeutics for gene knockdown and some new splice modulation opportunities. mdRNA, of course, is scheduled to run out of cash early next year and it would be a miracle if the two companies had not contemplated such a merger, particularly after AVI Biopharma moved in as mdRNA’s neighbor not too long along up from Oregon. With AVI’s cash reserve, the combined company may then find it easier to attract platform partners for mdRNA’s technology further bolstering the financials.

3) Silence Therapeutics and Archemix (not really an antisense company, but close enough). It is almost two months now that Silence announced to be in reverse merger talks. Since then, however, not a word except for so-so news on the results of opposition proceedings at the EPO related to their core patent. What Silence needs is cash (who doesn’t?), and what it can offer is a phase I cancer program, RNAi Therapeutics drug development expertise, and an siRNA structure that is not without use, although facing very serious patent challenges. What Archemix needs with its growing pipeline is access to public markets as evidenced by a previously failed reverse takeover attempt with cash-rich NitroMed, as does by the way Quark Pharmaceuticals which should also be counted as a possible Silence Therapeutics merger candidate. The fact that Silence Therapeutics has some aptamer in its blood line and the increased investment by Archemix into aptamer-mediated delivery of RNAi (Dicerna) and microRNA (miRagen) Therapeutics should help a combined company find a common language. Until any such deal is announced, however, Archemix would have to be prepared though that yet another potential partner will walk away from it last minute as Silence Therapeutics should also be receptive to other offers.

Which one is the most likely combination? Add your vote on the right.

Cequent Pharmaceuticals to Enter Clinic with First Oral RNAi Therapeutics

Cequent Pharmaceuticals is somewhat of an oddity in the RNAi Therapeutics space. Neither does it use synthetic RNAi triggers, nor does it fit properly into the DNA-directed RNAi paradigm. Instead, it is based on the so called trans-kingdom RNAi (tkRNAi) technology where genetically engineered bacteria are utilized to both generate and deliver the RNAi trigger, particularly to areas where commensal bacteria occur naturally. This includes the epithelia of the gastrointestinal tract, skin, and urogenital areas. This differentiation then allowed it to become the first RNAi Therapeutics company to test an orally administered RNAi Therapeutic in the clinic. The candidate drug, CEQ508, aims to prevent the formation and malignant progression of polyps in Familial Adenomatous Polyposis (FAP).

FAP is a rare ('orphan') inherited disease in which mutation of the tumor suppressor APC gene initiates events that cause the growth of countless polyps that ultimately progress to become malignant by the age of 40-50. CEQ508 contains an shRNA that is targeted against beta-catenin which is the major mediator of APC-regulated signaling, and the knockdown of which should therefore delay polyp proliferation and development of malignancies. In the trial, 18 adult FAP patients will be given capsules containing lyophilized tkRNAi bacteria daily for 28 days. Of course, safety is the primary objective of this phase I trial, but beta-catenin levels will also be measured as an early biomarker for drug efficacy. A cute video from Cequent's website explains the technological approach of this trial much better than I could ever do here:

Let me say a few general words about tkRNAi. When I saw the seminal paper initially, years before my direct interactions with the company (disclosure: I have been consulting for the company), I was somewhat skeptical of the approach. One of the reasons was that bacteria are very complex systems compared to other RNAi delivery systems, which means that there should be an increased likelihood for causing off-target phenotypes. The other concern was that maximal knockdown levels in vitro appeared to be less than typically achieved with siRNAs and shRNAs. Since then, however, I have learned a little bit more about tkRNAi convinced myself that it indeed does knock down genes, and that at this stage a carefully chosen target could allow for clinical success at ~50-60% knockdown levels as observed in the CEQ508 non-human primate studies. As to the off-target effects, the tkRNAi products have been shown to be well tolerated in a number of animal models. In addition, I learned that Cequent was not the first to harness bacteria as therapeutic agents for the transfer of genetic material so that additional safety and mechanistic information is available already.

Because of the awesome powers of bacterial genetics I would expect that the speed and creativity with which specificity (e.g. engineering of targeting ligands; reduction of key components responsible for off-targeting; increase of the ratio of shRNA to bacterial RNA and DNA following release into cytoplasm etc) and potency (e.g. shRNA designs; co-expression of factors affecting shRNA maturation etc) can be increased should be quite considerable. On the other hand, with the recent fund raising, it will be the immediate focus of the company to conduct the phase I trial where it will gain first clinical experience with the system in the gastrointestinal tract, lessons from which should also be also applicable towards follow-up gastrointestinal applications such as the inflammatory bowel disease program it co-develops with Novartis.

Combining Forces to Understand RNAi Therapeutics Inside Out

This entry is first a PR on a business development of the blog itself, but also reflects what should be the next major value-driver in RNAi Therapeutics after the liver: solid cancers.

Solid cancers are a very attractive target for RNAi Therapeutics because of its unmet needs and because of the so called Enhanced Permeability and Retention (EPR) effect of solid tumors which means that it should be possible to target them with siRNA-containing nanoparticles. Getting there, of course, is only half the story. The particles need to navigate their way through the extracellular matrix of tumors, latch onto the cancer cells, be taken up, and then finally be released into the cytoplasm. This requires a good understanding of nanoparticle-related chemistry and physiology, the biology of the extracellular matrix of cancer tissues, and cancer cell membrane biology.

Enter Tobias Wolfram. I have known Tobias since my days studying biology in Heidelberg (10 years ago now!) and have since been impressed by his enzyclopaedic knowledge of not only biology, but also history, psychology, economics and what not. He spent much of his time as a teenager with science projects and in molecular biology labs and won national prizes, at a time I did not even know that PCR existed. Since then he has become a truly multi-discliplinary scientist spanning the subjects of biology, chemistry, and material physics. Right now, he is at the Max-Planck-Institute for Metals Research in Stuttgart, the home of German engineering, where Tobias employs precisely engineered nanometer-patterned substrates for studying the interaction of cells with the extracellular matrix and their use for cell-based diagnostics. I visited him there two weeks ago to talk about working more closely together on the topic of RNAi trigger delivery.

In an experiment, we have decided to combine his expertise in getting molecules to cells with my understanding of the molecular biology of RNAi inside cells, to hopefully provide more insightful blog entries on the topic of RNAi Therapeutics delivery, with an initial emphasis on solid cancers. With Calando, Alnylam, and Silence Therapeutics having active programs in solid cancers, we will, over the next couple of weeks, start by taking a look at each of the applied technologies.