RONDEL Study Sheds Light on Fate of Cationic RNAi Delivery Systems

As regular readers may know already, I am not convinced that the polycation-based RONDEL delivery technology, and therefore Arrowhead’s CALAA-01 which is in clinical development for advanced solid cancers have bright prospects. In addition to the immune stimulation concerns which only came to the fore after the human RNAi proof-of-concept Nature paper (Davis et al. 2010) was published and which may be correctable if Arrowhead would modify their siRNAs, it is the limited pharmacokinetic profile with this delivery technology that makes it unlikely that the formulation can effectively take advantage of the enhanced permeability and retention (EPR) effect for tumor delivery. New research (Zuckerman et al., 2012) by the Davis group from Caltech, which is also the inventor of this technology, sheds light on the likely mechanism behind the pharmacokinetic profile. Importantly, this research has implications for the design of all delivery technologies involving complex formation between RNAi triggers and constitutively positively charged polymers and lipids (e.g. AtuPLEX, PEI-based polyplexes).

Previous biodistribution studies (Bartlett et al, 2007) showed that much of the RONDEL-delivered siRNA ended up in the kidney and bladder. At the time, it seemed plausible that this was due to displacement of the short siRNAs from the polycation backbone by competing negatively charged components in the blood. Thus, the pharmacokinetic behavior of the siRNAs would be almost like that of naked siRNAs which are known to be rapidly excreted through the kidneys. The new research, however, looked in more details at the pharmacokinetics of siRNA and delivery components in the kidney-bladder. This revealed that, in fact, the formulated siRNAs followed a slightly different profile than would be expected from that of naked siRNAs. Bringing to bear a number of sophisticated visualization technologies, it was found that the nanoparticles were first deposited intact in and around the glomerular basement membrane (GBM) before falling apart into their constituent components which, in turn, were small enough to escape through the slit pores into the urine (intact nanoparticles would be too large to make it into the urine).

Further adding credence to the notion that the RONDEL particles remain intact in circulation was a report (Oney et al., 2009) that the polycation backbone chemistry in RONDEL can be used to scavenge free oligonucleotides in circulation, which can be useful e.g. to regulate the activity of aptamer therapeutics. Similarly, the research by Zuckerman et al. shows that mixing separately added siRNA and RONDEL components in blood allows for the reconstitution of the nanoparticles.

The reason for the instability in the GBM is thought to be due to the strong negative charge of this extracellular matrix, meaning that the siRNAs are competed off the polymers there. I was quite surprised to learn during my cursory review of GBM biology that cationic polymers such as polyethyleneimines (PEI), a common nucleic acid delivery chemistry, are used to visualize the GBM! It is therefore likely that what was found for RONDEL will have implications for the pharmacokinetics, and safety, of other cation-based delivery systems. I mention safety, too, since it is known that particulate deposits (e.g. antibody-antigen) in the GBM can often cause glomerulonephritis (‘leaky kidney’). Whether the latter will be a problem may also depend on the duration the complexes reside there. At least for RONDEL, the time seems to be quite limited (on the order of minutes).

Preclinical validation of CALAA-01 in tissue culture and rodent studies

In addition to the pharmacokinetic study by Zuckerman and colleagues, another CALAA-01-related study was just published (Rahman et al., 2012). Conducted in Atlanta, with help from Caltech, this research is more optimistic about CALAA-01 by adding data in support of the validity of RRM2 as a cancer target. I myself have had problems understanding why this gene was chosen in particular when there are so many other attractive targets out there for therapeutic cancer RNAi. At least the tissue culture data strongly suggest that there is a target-dependent inhibition of cell proliferation. The animal knockdown data were not as strong, but some target knockdown, and even more pronounced anti-cancer effects were observed. Unfortunately, this being with a non-modified siRNA, one may want to take this with a grain of salt.

RNAi Therapeutics Financial Viability Looking Up Following String of Clinical Results

By: Dirk Haussecker

Note: A PDF version of this article is available at myfirstnameDOTmylastnameATgmailDOTcom

Abstract

Shortly after the 2006-8 period of exuberance during which access to capital was easy, the RNAi Therapeutics industry found itself in a financially difficult position. At the roots of this change were the eventual recognition of some poor science, clinical setbacks, and the tension arising from the more gradual progress of science and impatient markets. Clinical validation of RNAi-mediated gene knockdown following systemic delivery was seen as the only way out of this situation. This review summarizes how such critical validation was provided by a series of recent clinical results from the ALN-VSP02, Atu027, ALN-TTR01, and ALN-PCS02 development programs. These results are expected to reinvigorate investments in the technology.

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Background

In the spring of 2011, the RNAi Therapeutics industry had just gone through two financially very stressful years. A few large pharmaceutical companies (‘Big Pharma’), among them Roche and Pfizer, made high-profile exits from the space [1], pure-play RNAi Therapeutics companies were crashing, and optimism gave way to a sense that RNAi in humans may take much longer to achieve than once thought due to the delivery challenge and the risk of causing immune stimulations [2]. This negative view of the technology and its financial prospects came sudden. After all, it was only in 2006-8 that the medical importance of RNAi had been recognized with the Nobel Prize in Physiology or Medicine [3], and Merck and Roche alone accounted for almost $2B of investments in the technology.

The argument can be made that the RNAi Depression was catalyzed by the US Housing Crisis and the ensuing global financial turmoil towards the end of 2008. This triggered a retrenchment of capital from high-risk innovation, capital that failed to return to the RNAi space. The seeds, however, had already been planted during the RNAi Therapeutics hype phase. As often is the case in such periods, it attracted the attention of the notorious promotional biotech schemes which in turn were readily followed by the fast money crowd, all of which, of course, did not mix well with the more gradual pace of technological progress. The scientifically leading pure-play RNAi Therapeutics companies can be accused as well for having catered to the cravings of fast money by over-promising on timelines and pushing programs into the clinic which were not adequately mechanistically validated (e.g. clinical candidates based on the local administration of naked, sometimes entirely unmodified siRNAs to the eye and respiratory epithelium). Similarly, the scientific community and journals can be blamed for failing to better police the quality of the science that got published [4,5]. Consequently, considerable investments were made (and wasted) with little discernment to what was deserving science and what was not. When it was realized that only a very few of the claimed technical solutions had clinical potential, the equally erroneously conclusion was drawn that the technology as a whole had poor prospects instead of realizing that only a very few successful platform technologies can already create considerable value.

With investors tiring of ‘promising’ pre-clinical results and refusing to put more money into RNAi Therapeutics development unless clinical validation was seen, it was up to the science to prove itself in Man. If not, probably all of the significant pure-play RNAi Therapeutics companies, possibly with the exception of Alnylam, would find it impossible to raise capital on acceptable terms. Fortunately, despite the attendant contraction in the number of new development programs, some of the early capital meant that four candidates in particular had entered clinical development in time to be the industry’s chance at unambiguously validating RNAi in Man: ALN-VSP02, Atu027 ALN-TTR01, and ALN-PCS02. If this could be achieved, it was likely that capital would return to the space. If not, the added wait before such clinical validation could come would have been a great setback to the industry, a setback from which it might have been difficult to recover from financially.

ALN-VSP02 and Atu027: clinical safety of two leading delivery technologies

Together with CALAA-01 (sponsor: Arrowhead Research), Alnylam’s ALN-VSP02 and Atu027 by Silence Therapeutics were the three leading RNAi Therapeutics candidates in cancer, with enrolment starting in 2008-2009. Not only were these candidates important in their own right for their medical and commercial potential, they were supposed to clinically validate the three distinct systemic delivery platforms on which they were based: the cyclodextrin-containing polycation RONDEL technology (CALAA-01), the AtuPLEX lipoplex technology (Atu027), and SNALP liposomes (ALN-VSP02). In addition to cancer, together with hepatic applications the commercially most critical application of RNAi Therapeutics in the near to medium term, AtuPLEX has potential for endothelial cell-directed gene knockdown in general, and systemically administered SNALP also for liver and phagocytic cell-directed gene knockdown.

CALAA-01 enjoyed a year head-start and reported a first data update in a high-profile paper in April of 2010 [6]. Attracting the widespread interest was the fact that by taking tumor biopsies, the investigators were able to demonstrate, with the help of the 5’ RACE assay on tumor biopsies, that RNAi had occurred in target tissues. Moreover, evidence was provided that consistent with this functional finding, RONDEL nanoparticles could be detected in the biopsies. On the other hand, the data on the target knockdown was more ambiguous. Although the RNA analysis suggested measurable gene suppression, the protein analysis did not fully support that. The early promise was only short-lived, however, as a subsequent 2010 ASCO presentation (Abstract No: 3022) showed ample innate immune stimulations which forced the company to concede in late 2011 that at least another phase I trial was necessary in an effort to better manage them. In retrospect, it seems obvious that a major omission of the program was in leaving the RNAi trigger (targeting the M2 subunit of ribonucleotide reductase, RRM2) chemically unmodified, meaning that the risk of inducing such responses was quite high. In the absence of evidence for clinical efficacy and unexplained trial delays (it has taken ~3 ½ years to conclude enrolment), the CALAA-01 phase I results were thus unable to positively impact perceptions of RNAi Therapeutics.

An important step forward in that direction was made with the 2011 ASCO presentations of the fully enrolled phase I study of ALN-VSP02 and a quite encouraging interim update for Atu027. The data suggested that Atu027, an endothelial cell-directed multilamellar cationic lipoplex containing an RNAi trigger against PKN3 [7], was surprisingly well tolerated as dose escalation had reached dosages at which gene knockdown efficacy could be expected based on the preclinical animal data. Dose escalation has been ongoing since and has exceeded predicted RNAi-functional doses- and still no dose-limiting toxicities or serious adverse events were claimed as of December 2011. The reason why this is somewhat unexpected is that the positively charged lipoplexes may have been considered prone to induce various innate immune responses [8], also because no immune suppressive regime was used in the trial. It is possible that the extensive 2’-O-methylation of the AtuRNAi-type trigger partly accounted for that. Activations of the alternative complement pathway, however, were noted, although these were claimed to be clinically not significant. Tumor responses by stringent RECIST criteria meanwhile remain to be demonstrated. Nevertheless, the overall safety profile and well-behaved pharmacokinetics have encouraged further investments in the AtuPLEX and related cationic lipoplex delivery platforms from Silence Therapeutics and can be considered a meaningful step forward for the field.

Of the three candidates, the most ambitious phase I study was that for ALN-VSP02, a SNALP formulation that includes two siRNAs, one against VEGF and KSP for anti-angiogenic and anti-proliferative mechanisms of action, respectively. Enrolling 41 patients with advanced solid cancer with liver involvement, this study included a battery of tests, including biopsies to test for target mRNA cleavage and siRNA tissue concentrations, the measurement of tumor blood perfusion as an indicator of anti-VEGF activity, and various other pharmacological parameters. The study succeeded in demonstrating that this SNALP formulation was fairly well tolerated in these advanced cancer patients at dosages of up to 1.0-1.25mg/kg (ASCO 2011 poster #3025). Among the dose-limiting toxicities were a liver failure with subsequent death at 0.7mg/kg, two cases of transient grade 3 thrombocytopenia at 1.25mg/kg, a grade 3 hypokalemia at 1.5mg/kg, and four grade 1-2 rigor/chills at 1.0mg/kg (one case) and 1.25mg/kg (three cases). The death at the 0.7mg/kg dose was deemed to be possibly related to study drug and occurred in a patient where the tumor burden in the liver was quite extensive. The enrolment criteria were subsequently adjusted to exclude similar patients with a greater than 50% tumor burden in the liver.

Based on the pre-clinical studies (AACR 2009 poster #B204), knockdown activity could have been expected at dosages of 1.0-1.25mg/kg. Consistent with this notion, the 5’ RACE for VEGF (but not KSP) RNAi cleavage performed on the biopsies revealed RNAi activity. In terms of functional activity, the Ktrans, a measure of tumor blood perfusion, declined by 40% or more in over half the patients that had received one or more doses and was consistent with anti-angiogenic activity of the drug. It has to be said though that the Ktrans response was not nearly dose related. Evidence for dose-responsiveness, however, was provided when categorizing patients according to RECIST criteria of tumor responses. Accordingly, only one in 13 patients treated with up to 0.4mg/kg had stable disease for two months or more. This compared to 12 of the 24 given higher doses. Moreover, there was a 70% partial tumor response in an endometrial cancer patient at 0.7mg/kg who stayed on study drug for months; at the 1.0mg/kg dose, the recommended dose for further studies, 7 of 11 achieved stable disease. Despite these preliminary signs of activity, there remain questions about the choice of the target genes, particularly the suitability of VEGF as an RNAi target and the absence of detectable KSP cleavage. Moreover, systemic SNALP delivery to even tumors in the liver is thought to require extended blood circulation times in order to harness the EPR effect, yet the half-life of siRNA in the blood was only in the 15-30 minute range. This is consistent with the relatively short C14-PEG lipid anchor in this formulation. Notwithstanding, the ALN-VSP02 study was a stringent test for the safety of SNALP delivery and added considerable clinical pharmacokinetic experience to this technology. Among the latter was the detection of amounts of siRNAs in normal liver which strongly indicated [9] that gene knockdown with SNALP was possible for liver-expressed genes.

ALN-TTR01 and ALN-PCS02: most impressive demonstrations of RNAi in Man

Despite the largely acceptable safety and promising pharmacokinetic data from the Atu027 and ALN-VSP02 studies, the field still lacked black-and-white evidence for target gene knockdown following systemic delivery. The SNALP-enabled TKM-ApoB by Tekmira in early 2010 was close to providing such evidence. Unfortunately, concomitant with the achievement of slight ~20% ApoB reductions, moderate immune stimulations were observed at 0.6mg/kg in the dose escalation trial, causing the company to terminate the trial (http://clinicaltrials.gov/ct2/show/NCT00927459?). The burden thus fell onto ALN-TTR01, another SNALP-enabled RNAi Therapeutics targeting the liver-expressed transthyretin gene. Mutations of this genes frequently cause familial amyloidotic polyneuropathies and cardiomyopathies which shorten the lives of ca. 50,000 patients worldwide.

This time implementing transient immune suppression (corticosteroids and H1/H2 histamine receptor blockade) as a precautionary measure, the trial began enrolling patients in June 2010 in Europe. A little more than a year thereafter, Alnylam presented almost full trial data at the November 2011 FAP conference in Kumamoto, Japan. It came as a great relief that at the highest, 1.0mg/kg dose the 5 patients exhibited a mean reduction of serum TTR of 41% following a single intravenous infusion. One patient exhibited a text-book 81% RNAi-type TTR reduction at nadir (week 1), with pronounced knockdown persisting out to 4 weeks (50%). Without ifs or buts, this was clear demonstration of effective RNAi in Man. Equally important, except for mild-to-moderate infusion reactions which were readily managed by simply slowing the rate of infusion, no meaningful adverse events were reported. As that trial is currently being wrapped up with more patient data expected for the 1.0mg/kg dose cohort, the sponsor Alnylam is aiming to further enhance the competitive profile of its TTR candidate and has filed a CTA for clinical trials with a new, ALN-PCS02-type SNALP formulation for which equivalent gene knockdown can be expected at 10-fold or more reduced dosages (patent application WO 2010/144740 A1). The goal here is to achieve a more potent knockdown with a higher margin of safety and a once every month or two dosing frequency.

The RNAi clinical dataflow culminated in early January 2012 with Alnylam announcing dose escalation data for the phase I study of ALN-PCS02 for the treatment of hypercholesterolemia. As predicted, equivalent knockdown to ALN-TTR01 were obtained at much reduced dosages with this improved ‘MC3-type’ SNALP formulation from Tekmira: ~60% mean peak reductions in serum PCSK9 for the 0.15mg/kg and 0.25mg/kg dose cohorts. As PCSK9 antagonizes ‘bad’ LDL cholesterol removal from circulation [10], its inhibition was accompanied by ~35% reductions in LDL cholesterol. Due to the favorable safety profile (rashes were noted, but these were likely related to route of administration as they also occurred in the placebo cohort), dose escalation is expected to proceed. At the higher dosages, more robust, less variable knockdown can be expected for both PCSK9 and LDL-cholesterol. This would put it in a favorable competitive position vis-à-vis the PCSK9 monoclonal antibody competition (e.g. REGN727/SAR236553).

The most significant limitation of the ALN-PCS02 study turned out to be the use of transient immune suppression. These caused short-lived (+65%) spikes and (-25%) depressions of PCSK9 and LDL-cholesterol, respectively. While transient immune suppression should be acceptable for many of the initial indications of high unmet medical need for which SNALP-based therapeutics are largely being developed at the moment, especially if once every month or two dosing can be achieved, they can not only complicate the analysis of studies such as ALN-PCS02, but would also restrict the eligible patient population for a condition such as hypercholesterolemia. Introduced as a precautionary measure following the TKM-ApoB experience, the question is whether it is possible to do entirely without it as SNALP potency has improved [11], lipid-specific toxicities been minimized, and more predictable innate immune stimulation assays been introduced as presented by Tekmira at a Drug Information Association (DIA) meeting on March 23, 2010, in Bethesda, MD.

Conclusion

The clinical results not only provided the long-awaited clinical validation and are a boost of confidence for the entire RNAi Therapeutics industry, but they directly de-risk two of the most important systemic delivery technologies: SNALP and AtuPLEX. The results with SNALP in particular set the stage for a forceful expansion of this delivery platform with already 5-6 candidates in active clinical development: ALN-VSP02, ALN-TTR01+02, TKM-PLK1, TKM-EBOLA, and ALN-PCS02. More still are expected to enter the clinic over the next two years. The following months should also add to the SNALP clinical experience in the form of results from the fully enrolled and dose-escalated ALN-TTR01 and ALN-PCS02 trials, an update on Tekmira’s cancer therapeutic candidate TKM-PLK1, and safety data from the TKM-EBOLA volunteer study. Since the ASCO 2011 presentation, Atu027 has attracted commercial interest in the form of various technology evaluations of AtuPLEX and related delivery technologies from Silence Therapeutics (partners: InteRNA Technologies, Mirna Therapeutics, an undisclosed Japanese ‘Top Ten’ global pharmaceutical company, and miRagen).

The space, however, awaits confidence expressed in the form of a more major financial commitment by a larger pharmaceutical company. This might break the gridlock caused by Roche’s decision to stop in-house RNAi Therapeutics development, a decision so powerful that it essentially caused all capital to retrench to the sidelines or leave RNAi Therapeutics entirely. What should not be lost is that the current situation also represents an attractive technical risk-financial reward opportunity for those companies that dare rely on their own scientific instincts rather than follow the herd. This includes mid-sized pharmaceutical companies and those in the newly emerged and emerging economies which are playing an increasingly important role in RNAi Therapeutics. As access to capital normalizes, platforms in addition to SNALP and AtuPLEX will likely emerge and help further expand the therapeutic reach of RNAi Therapeutics.

Abbreviations

5’ RACE: 5’ rapid amplification of cDNA ends; Ktrans: volume transfer coefficient; RECIST: Response Evaluation Criteria of Solid Tumors; RONDEL: RNAi/Oligonucleotide Nanoparticle Delivery; siRNA: small interfering RNA; SNALP: stable nucleic acid lipid particle.

References

1. Ledford H: Drug giants turn their backs on RNA interference. Nature 2010, 468: 487.
2. Robbins M, Judge A, Ambegia E, Choi C, Yaworski E, Palmer L, McClintock K, MacLachlan I: Misinterpreting the therapeutic effects of small interfering RNA caused by immune stimulation. Hum Gene Ther 2008, 19: 991-999.
3. Zamore PD: RNA interference: big applause for silencing in Stockholm. Cell 2006, 127: 1083-1086.
4. Tolentino MJ, Brucker AJ, Fosnot J, Ying GS, Wu IH, Malik, Wan S, Reich SJ: Intravitreal injection of vascular endothelial growth factor small interfering RNA inhibits growth and leakage in a nonhuman primate, laser-induced model of choroidal neovascularization. Retina 2004, 24: 132-138.
5. Aouadi M, Tesz GJ, Nicoloro SM, Wang M, Chouinard M, Soto E, Ostroff GR, Czech MP: Orally delivered siRNA targeting macrophage Map4k4 suppresses systemic inflammation. Nature 2009, 458: 1180-1184.
6. Davis ME, Zuckerman JE, Choi CH, Seligson D, Tolcher A, Alabi CA, Yen Y, Heidel JD, Ribas A: Evidence of RNAi in humans from systemically administered siRNA via targeted nanoparticles. Nature 2010, 464: 1067-1070.
7. Aleku M, Schulz P, Keil O, Santel A, Schaeper U, Dieckhoff B, Janke O, Erdruschat J, Durieux B, Roeder N, Löffler K, Lange C, Fechtner M, Möpert K, Fisch G, Dames S, Arnold W, Jochims K, Giese K, Wiedenmann B, Scholz A, Kaufmann J: Atu027, a liposomal small interfering RNA formulation targeting protein kinase N3, inhibits cancer progression. Cancer Res 2008, 68: 9788-9798.
8. Yew NS, Scheule RK: Toxicity of Cationic Lipid-DNA Complexes. Adv Genet 2005, 53PA: 189-214.
9. Landesman Y, Syrzikapa N, Cognetta A 3^rd, Zhang X, Bettencourt BR, Kuchimanchi S, Dufault K, Shaikkh S, Gioia M, Akinc A, Hutabarat R, Meyers R: In vivo quantification of formulated and chemically modified small interfering RNA by heating-in-Triton quantitative reverse transcription polymerase chain reaction (HIT qRT-PCR). Silence 2010, 1: 16.
10. Horton JD, Cohen JC, Hobbs HH: Molecular biology of PCSK9: its role in LDL metabolism. Trends Biochem Sci 2007, 32: 71-77.
11. Semple SC, Akinc A, Chen J, Sandhu AP, Mui BL, Cho CK, Sah DW, Stebbing D, Crosley EJ, Yaworski E, Hafez IM, Dorkin JR, Qin J, Lam K, Rajeev KG, Wong KF, Jeffs LB, Nechev L, Eisenhardt ML, Jayaraman M, Kazem M, Maier MA, Srinivasulu M, Weinstein MJ, Chen Q, Alvarez R, Barros SA, De S, Klimuk SK, Borland T, Kosovrasti V, Cantley WL, Tam YK, Manoharan M, Ciufolini MA, Tracy MA, de Fougerolles A, MacLachlan I, Cullis PR, Madden TD, Hope MJ: Rational design of cationic lipids for siRNA delivery. Nat Biotechnol 2010, 28: 172-176.

A Very Obvious Method of Generating Tuschl-type siRNAs

John Leavitt over at the RNAi Litigation blog just posted ‘Alnylam’s’ defense against the accusation of the University of Utah that Utah had been deprived of ownership over the 3’ overhang feature of the Tuschl-type siRNAs. Setting aside the underlying merits of the case, the Motion to Dismiss the Second Amended Complaint by the University of Utah highlights one thorny issue with the granted Tuschl II (T-II) patents in the US, namely that they, so far, do not claim the 3’ overhangs themselves, but a method of generating 3’ overhang siRNAs. This method stipulates, in a first step, the synthesis of the individual strands, and then, in a second step, hybridizing (‘combining’) them to form the 3’ overhung siRNAs. The motion by the Defense consequently argues that Utah is missing the point in its suit by alleging ownership over the 3’ overhang feature, but not the method of generating 3’ overhung siRNAs subject of the US patents.

This to me is a) misleading since the Tuschl II IP estate to which Utah lays claim includes the European T-II patent which expressly claims the overhangs and Alnylam/Max Planck are obviously hoping to get similar composition of matter claims issued in the US, and b) the issued US T-II could only be considered novel by the USPTO based on the novelty and utility of the composition of matter that results from this method, i.e. 3’ overhang siRNAs for inducing gene silencing.

That the USPTO issued these claims in the first place is quite puzzling. Synthesizing and combining small RNAs is the most obvious method of generating 3’ overhung siRNAs. I would argue that even the average highschool student can come up with this method after a basic lesson on nucleic acid structure, not to speak of the ‘person having ordinary skill in the art’ which is considered the standard for obviousness. Unless I have missed an important exception in US patent law, similar to the Swiss-style claim construction in Europe to which the US T-II claims are reminiscent of, the US T-II claims seem very tenuous to me and probably should not have been granted.

It is amazing that both Plaintiffs and Defense are spending all this energy (=time and legal fees) skirting around the main issues (namely that Tuschl was probably motivated to test the 3' overhang feature based on Bass' speculations, and that Bass cannot be named a (co-)inventor since there was not even a semi-formal collaboration between Bass and Tuschl and Utah never bothered to file a patent). The answer by Utah to this motion is predictable, and so it will go on and on...

Here Comes the Stock Offer!

After all the brown-nosing* by the analyst and investment bank community, Alnylam just announced that it has commenced a public offering of 7,000,000 shares. This should net the company around $85,000,000- probably enough for a low-ball offer for Tekmira.

Yes, Alnylam is burning through its cash at a rate of $80M a year, and yes, it is an old adage in biotechnology that you raise capital when the going (= share price) is good. With $260M in the coffers though, the timing would seem overly conservative and given my read on the situation, it is in Alnylam’s best interest to make Tekmira an offer. Valentine’s Day would be good timing for that indeed.

* I’ll wash my finger-tips, but I had to get this off my chest. There are analysts that have been there through good and bad times. Others have understandably been attracted by the positive clinical trial results- OK. But the recent analyst upgrades and Q+A sessions in conference calls were just unbearable with nothing but softballs and the avoidance of tough questions.

The Business of RNAi Therapeutics in 2012

As I tweeted already, a new journal devoted to nucleic acid-based therapeutics, Molecular Therapy-Nucleic Acids (MTNA), just came online. The first articles already indicate that this journal is poised to become an important addition to the field of RNAi Therapeutics: promising siRNA chemistry work by Merck, folate-siRNA conjugates by Roche, and an article on SNALP and 'lipidoid' delivery to immune cells by Alnylam. Today, a review by myself on 'The Business of RNAi Therapeutics in 2012' analyzes the dramatic ups and downs this field has experienced in its less than 10 years in existence, with a particular focus on the downturn following the 2007/2008 peak. I believe that it is timely in that the industry is entering a new, recovery phase.

If you are interested, an Open Access version of the article is available here.

Alnylam Squares Off with Dicerna

Dicer-substrate RNAi triggers are often seen as a (probably cheaper) alternative to Tuschl siRNAs. Initially, based on a small sample size, it was even claimed that Dicer-substrates had superior potencies and prolonged durations of knockdown. Of course, Alnylam, considering Tuschl siRNAs to be its property, has recognized this and regards Dicer-substrates along with its corporate champion, Dicerna, a competitive threat. Although Alnylam has long claimed that Tuschl siRNAs are preferable over Dicer-substrates for various reasons, until now it has largely been Dicerna’s word against Alnylam’s word.

This has changed with a publication by Alnylam in the journal RNA which provides a comprehensive, and I believe fair comparison between the two structures (Foster et al 2012). Comparing large numbers of RNAi triggers both in vitro and in KC2-SNALP animal studies, the study shows that Tuschl siRNAs and Dicer-substrates are essentially equivalent in terms of potency and the duration of knockdown. However, when these structures are compared in terms of innate immune stimulation, Tuschl siRNAs had a very slight edge when unmodified sequences were tested. Of course, it is well recognized that chemical modifications are required and very effective at abrogating these immune responses. When these were applied, the potencies of Dicer-substrates were more likely to suffer than those of Tuschl siRNAs, and in a few cases innate immune stimulation was not entirely abrogated. The former can be explained by the additional requirement for the Dicer processing step which can be affected by chemical modification.

Overall, this means that it may take a little bit more effort to identify a suitable Dicer-substrate clinical development candidates, and there could be an increased risk in encountering unforeseen innate immune stimulations in humans. On the other hand, the study also suggests that for some genes it may be possible to find more potent RNAi triggers with Dicer-substrates, so that in an ideal world one would keep an open mind. I should also add that, not discussed in this paper, there are also other considerations which may favor one structure over the other.

Unfortunately, we are not living in an ideal RNAi Therapeutics world, but one in which patent trolls and IP freeloaders abound. The timing of the comparison study is particularly ironic since the freedom-to-operate of US-based Alnylam is very much in doubt, thus increasing the attractiveness of Dicerna's offering. This is because of the recent issuance of the Baulcombe patent in the US which has put essentially all the Tuschl siRNA structures, bar one, in a straitjacket. Until Baulcombe is sorted out, all buy-out and partnering efforts will be on hold, and if Merck gets exclusive rights to that patent...then Alnylam may well be toast.