Friday, December 30, 2011

Dicerna Ends Year on Positive Note

With the Notice of Allowance of a US patent application covering its basic Dicer-substrate RNAi trigger technology and a $5M milestone payment from Kyowa Hakko Kirin (KHK) due to the formal initiation of development work for a Dicer-substrate oncology candidate by its Japanese partner, Dicerna can look optimistically into the New Year. These events also symbolize some of the important shifts that have taken place over the last 3 years, namely the increasing choice between RNAi trigger technologies and the increasing importance of Asia in the industry.

Dicer-substrate patent

The patent that is about to formally issue covers methods related to knocking down genes in mammalian cells using certain 25-30 base-pair double-stranded RNAs that undergo Dicer processing before incorporation into the RISC gene silencing complex and target cleavage. While not gate-keeping for Dicer-substrates, the patent will give Dicerna considerable clout over this particular gene silencing approach and has an expiration of at least 2025 if not a year or two longer following patent term adjustments. It is interesting to speculate that the Allowance was also a trigger for the $5M milestone payment.

New oncology candidate formulated in KHK delivery tech

The press release on the $5M milestone noted that while the RNAi trigger of the candidate that is at the center of the $5M milestone payment is licensed from Dicerna (possibly targeting KLF5), the drug delivery tech would be KHK’s own. Based on comments made in early 2010 when Dicerna and KHK first announced their collaboration plus quite a few liposome-related patent apps by KHK, it is likely that the delivery tech relates to a cationic liposomal formulation. I would be a bit wary though about the novelty and strength of these liposomes since at least the patent apps merely claim cationic LNP formulations of chemical compositions that have been known for years in the art an manufacturing method that may be somewhat unique to KHK, but maybe too complex for industrial applicability. Moreover, the described approach lacked the scientific rigor that I would expect of a company that intends to enter clinical development soon.

Dicerna employs Tekmira SNALP delivery

Dicerna has also been saying to be developing delivery technologies, including LNPs and another (actively) targeted delivery system. Interestingly, in an analytical assay development paper that has just been published (Jiao et al., 2011), Dicerna employed Tekmira’s SNALP delivery technology. It is unclear whether the SNALPs were reproduced based on the literature or whether there has been some sort of collaboration between the two companies. However, since Tekmira has been investigating a variety of non-Alnylam RNAi trigger technologies, the latter hypothesis is not that farfetched. The Allowance should provide further impetus for the two companies to work together more closely.

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Wednesday, December 28, 2011

Gradalis Swiftly Moves ddRNAi-Enhanced Cancer Vaccine Candidate through Clinic

I’ve been reminded a number of times by the staunch Benitec-supporters here that Texas-based biotech company Gradalis has been moving a ddRNAi-enhanced cancer vaccine candidate (‘FANG’) aggressively through clinical development. Virtually out of nowhere, Gradalis initiated clinical trials two years ago and there are now two active phase II trials, one in ovarian cancer and one for advanced melanoma. A peer-reviewed publication on the phase I trial was also just published (Senzer et al.) arguably making FANG the lead RNAi candidate in oncology.

The phase I study involved over 40 patients with advanced solid tumors and demonstrated the safety and logistic feasibility of the approach. Although evidence of suggestive of efficacy was presented such as a clear correlation between an immune response and survival, it would be premature to conclude anything with regard to efficacy. Having now followed a number of cancer vaccines, most of which have eventually failed, it seems to me that correlations such as this could be just as well as a reflection of the fact that those with more responsive immune systems will do better anyway.

FANG’ comprises of plasmid DNA from which a single RNA polymerase II promoter drives the expression of an upstream GM-CSF open-reading-frame followed by a pair of downstream RNAi hairpins. This plasmid is introduced by electroporation in a petri dish into the patients’ own cancer cells which have been obtained from a tumor resection. After allowing some time for the expression of the transgenes, the cells are irradiated so as to kill off their proliferative potential and are then re-introduced like many other vaccines by intradermal injection.

The GM-CSF component, wildly popular in the cancer vaccine field and also part of Dendreon’s famous prostate cancer vaccine PROVENGE, is supposed to serve as an attractant, proliferation and maturation factor for dendritic cells which are supposed to ingest, present and thereby stimulate an immune response against the antigens unique to a tumor; the pair of ‘bifunctional’ hairpins meanwhile both target furin which is thought to be an important protease for the maturation of the various isoforms of TGF-beta, a well-known immunosuppressant often overexpressed in cancer.

‘Bifunctional’ here means that one hairpin is perfectly matched and therefore mostly relies on the so-called Ago2/cleavage-dependent mode of RISC activation, whereas the other hairpin contains a central bulge due to mismatching changes introduced in the passenger strand arm of the hairpin thus relying on the non-cleavage pathway of RISC activation which can be facilitated by all four human Argonautes (both predicted to yield the identical guide strand). This strategy of distributing the RNAi between the various Argonaute proteins is certainly an interesting idea, but I’m not sure whether even Gradalis knows what consequences of this is both in terms of efficacy and safety.

A general lack of detailed molecular mechanistic studies is probably my biggest concern with this candidate and when thinking about Gradalis in general. It also at least partly explains why FANG has been moving so rapidly through the clinic. I find it particularly troubling that I have seen no detailed studies by Gradalis looking at the relationship between furin knockdown and TGFbeta inhibition which is key for Gradalis' strategy. This already has caused difficulties in interpreting some of the phase I data where possible assay problems complicated reconciling apparently only modest reductions in furin with much more pronounced down-regulations of TGFbeta. This not only makes it more difficult to make the right development decisions, but also when it comes to finding a partner for the program. On the other hand, you could argue that a cancer vaccine candidate involving both GM-CSF expression and TGFbeta inhibition already has a good chance at succeeding, and sweating out the technical details would only cause delays without making us much the wiser.

As I had mentioned, the Benitec supporters are following Gradalis’ development with much interest as such an advanced ddRNAi candidate may be a prime licensing opportunity for Benitec which controls an important part of the ddRNAi patent landscape. I’ve certainly looked at the hairpin structures involved in light of Benitec’s patent claims (esp. the ‘099 Graham patents) and there is a good chance that Gradalis ought to take a license as it further monetizes this candidate, although their structures may give them a bit of wiggle room.

Mirna Therapeutics selects Marina Biotech’s SMARTICLE delivery tech

In another notable development last week, cancer microRNA Therapeutics company Mirna Therapeutics said that it would use Marina Bio’s SMARTICLE liposomal delivery technology for the development of microRNA replacement therapy for cancer. Based on conference presentations, the two companies had been collaborating on the delivery of microRNA mimics before announcing the deal. An attraction of the SMARTICLE delivery technology, which Marina had acquired from Novosom, is certainly the fact that there is already clinical experience after SMARTICLE-enabled ‘DNAi’ compound by ProNAi has begun dosing a year ago. Similar to related agreements between Mirna and Silence, and InteRNA and Silence, insightful details about the financials were not disclosed. For Marina, which have diluted shareholders by what seems like a 100-fold over the last 3 years (unreal, really), it is good news as its extensive technology offering is finally getting takers.

Tuesday, December 13, 2011

Mr. Anonymous Thwarted in Zamore US Patent Re-exam

Silence Therapeutics reported that a number of valuable RNAi trigger patents related to the Zamore Design Rules that were issued last year in the US were upheld following a re-examination request by an anonymous 3rd party. Even stronger claims related to the same Zamore patent series have been issued in Europe, and unsurprisingly Alnylam, but also Novartis and Alcon are opposing them with the outcome to be decided (EP 1633890 B1). However, given the importance of both the US and Europe in the commercialization of innovative drugs, having a patent position in one jurisdiction alone can be considered a valuable strategic asset already.

The Zamore Design Rule patents are owned by the University of Massachusetts and exclusively licensed to Silence Therapeutics for medical uses. As described in more detail in other blog entries before (here and here), they cover methods of promoting the incorporation of the desired guide strand into the RISC gene silencing complex as well as of enhancing RISC turnover resulting in more effective and selective RNAi knockdown.

Especially the patent covering guide strand selectivity methods (‘thermodynamic end-stability rule’; US 7,750,144) is widely recognized in the art for greatly increasing the likelihood of finding efficacious RNAi triggers and is incorporated in essentially all bioinformatic sequence pre-selection algorithms. It has to be said though that the claims do not cover the entire spectrum of approaches of achieving differential end-stability. They do, however, cover chemical and structural approaches that have been reported by a few companies such as Marina Biotech and Sirna/Merck before. Moreover, because the coverage involves modified nucleotides, there will be the concern that even if such modifications were applied for other purposes (e.g. stability or immune abrogation), they may fall under the patents. Consequently, a company with a promising late-stage candidate may want to take a license instead of taking a chance in an infringement lawsuit.

This makes it the second time within a week (see PKN3 opposition by Alnylam) that important patents by Silence Therapeutics were upheld essentially unchanged following challenges. Alnylam’s management once laughed off the value of the Zamore patents in a conference call following their issuance last year. They obviously considered them serious enough to oppose them in Europe and Alnylam is certainly the most likely identity behind Mr. Anonymous (have your say by participating in the survey on the right). Less likely, but not entirely out of the question would be a Big Pharma company like Novartis which is considering taking a license to the Zamore patents, but first wanted to kick the tires before it did so (patents that have been unsuccessfully challenged are considered stronger).

With all these patent successes, and at least one more likely to come, Silence Therapeutics need to monetize their assets in the form of non-dilutive funding.

Hemophilia Gene Therapy Success Bodes well for ddRNAi Therapeutics

Following years of public scorn and derision, including by Alnylam which in 2006 waved off ddRNAi Therapeutics and gene therapy as ‘dangerous’, gene therapy is back with a vengeance. This week, a consortium of researchers reported in the New England Journal of Medicine that an self-complementary AAV8-delivered Factor IX transgene was able to significantly correct hemophilia B in a small clinical trial.

4 out of the 6 patients treated were able to largely discontinue the standard frequent (often 2-3 times a week) and expensive use of prophylactic recombinant FactorIX protein therapy which accounts for estimated healthcare costs north of $20M a lifetime. This means that even in the limited duration of the trial (6-16 months of follow-up), the low cost of the gene therapy ($30k cost of goods for a treatment that is expected to last many years if not a lifetime) meant that this therapy is already saving money (and improving quality of life). If the results can be confirmed in a larger trial and the side-effects, including transient liver enzyme elevations can be controlled with similar efficiency as in this small one, approval may not be that far away.

The hemophilia results also bode well for ddRNAi Therapeutics. First of all, the study has validated the safety and efficacy of delivering the highly promising AAV vector family by peripheral vein infusion to the liver. This represents progress over a previous hemophilia gene therapy study which employed considerably more invasive direct hepatic infusion. In particular, it is likely that, should the ddRNAi collaboration by Tacere/Pfizer for HepC continue, it would involve not only the same vector family (AAV) and target organ (liver), but also the same serotype (AAV8) and self-complementary genome strategy as employed in this hemophilia trial.

Saturday, December 10, 2011

Alnylam General Counsel Has Left Company

According to Linkedin, Alnylam's General Counsel, Philip Chase, has just left the company to become General Counsel of Adimab [note: an earlier version incorrectly stated that P. Chase had been with the company for only a little over a year, while in fact he had been there for 4 years]. This departure comes at a critical time for Alnylam as it is involved in a fierce battle with Tekmira for control over SNALP technology, and in which it has started to lose a number of preliminary battles (defamation suit and the City of Hope PR disaster). It also comes after various, possibly related, recent changes including the VP for Intellectual Property (Donna Ward), the lead attorneys in the VSP02 Interference case (Rothwell Figg), a CFO (Patty Allen), key business development personnel as well as two CSOs.

The strongmen at the helm of the company, the CEO and COO, and BoD, however, have largely remained intact during the upheavals over the past 2 years. In fact, John Maraganore (CEO) and Barry Greene (COO) were just re-'incentivized' in the form of options grants that will pocket them $150k and $100k for each US dollar increase in the company's share price (strike price near 3-year low, after 80% stock decline).

Alnylam program update

Based on a poster presentation at this year’s Amercian Society for Hematology meeting (ASH11), also the remaining target pick under Alnylam’s 5x15™ will be a SNALP-based program that targets a gene in the liver: Tmprss6 knockdown for the treatment of iron overload diseases. As such, all of Alnylam’s core clinical programs are SNALP-based. If you had looked at the pipeline 3 years ago, you would have thought that this was where Tekmira was heading. The challenge now will be to clinically mature the programs to a point (in 2015) where the package will be attractive enough for a company like Genzyme/Sanofi-Aventis to acquire the orphan drug company Alnylam for somewhere between $1.0-1.5B.

This development confirms signals that the company has recently started to send out (Rodman Renshaw investor note and comments in the Piper Jaffray presentation) namely that not only is Alnylam now advertising itself as a product, and not platform company, but that it is likely to be all about gene knockdown in the liver. Long gone seem the days when management predicted Alnylam to be the next Genentech. In light of the confrontation with Tekmira, this path opens up the possibility of a settlement whereby Alnylam is allowed continued access to SNALP for the 5 programs, with Tekmira getting full control over other SNALP applications in and especially outside the liver. As a product company that has committed on its programs, why bother with trying to control a platform which is giving Alnylam nothing but legal headaches.

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Friday, December 9, 2011

What Business Does Alnylam’s Counsel Have at the City of Hope?

The Tekmira and Alnylam dispute is a classical David versus Goliath encounter. By my standards, Protiva/Tekmira clearly deserves credit for developing the industry’s most advanced systemic siRNA delivery technology, SNALP. Alnylam, however, is trying to take all of that away from a company that it knows it stands a good chance at silencing with sheer money and influence.

It therefore struck a nerve when I read ‘Opposition 4’ in the ongoing VPS02-related patent interference filed on November 30 by Protiva/Tekmira (Interference No. 105792). It reveals that Alnylam’s counsel, Rothwell Figg*, went to the City of Hope, the employer of a key technical witness by Tekmira (John Rossi), to gently urge them that John Rossi should perhaps consider to withdraw his testimony (from Opposition 4):

For example, Material Fact 6, drafted by Alnylam’s counsel, clearly references “the meeting between representatives of City of Hope and Rothwell Figg.” Indeed, in questioning Dr. Rossi, Alnylam’s counsel appears to have acknowledged rather than disputed the occurrence of one or more meetings between Alnylam’s counsel and Dr. Rossi’s employer.’

And this is apparently what Dr. Rossi was informed of happened in those meeting(s):

‘[Dr. Rossi] Well, I was also informed that there was evidence that was evolved that would be used against my disclosure or my declaration in terms of the credibility of me as a -- basically as a scientist. In other words, I would be contradicting my own words, whether they be – I didn’t know what the source was of where my contradiction was going to be brought up, but it was in something that I had published or in publications or in patent applications or patents. That was all I was told. And I felt that it couldn’t be the case. I felt very strongly.

And in fact, Mr. Huntington asked me if I wasn’t worried about the fact that I could actually invalidate my own patents. I said no, not because I don’t want those to be invalidated, but because I didn’t feel that anything I had said or done would contradict those.’

It reads as if Alnylam’s counsel suggested to the employer of the witness that in following his conscience as a scientist, the employee would hurt the financial interest of his employer (City of Hope is the beneficiary Dr. Rossi’s patents). I’m no lawyer, and have no idea whether such apparent manipulation of the judicial process is legitimate or even legal, but it strikes me as very wrong (and desperate).

This episode though is just the latest in the game of politics Alnylam has been playing with Tekmira. It is high time for it to stop it and instead focus on the science. Otherwise, it risks wasting the opportunities that come with the ongoing clinical validation of Tekmira’s SNALP technology to which it is (still) a licensee.

* As I wrote in a blog entry in October, Rothwell Figg seem to have been replaced as Alnylam's counsel in the Interference, to be replaced by Wilmer, Cutler et al. which are also responsible for the larger SNALP Misappropriation case.

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Wednesday, December 7, 2011

SNALP RNAi versus RNaseH Antisense for Gene Knockdown in the Liver

Following recent phase I results from ISIS Pharmaceutical’s Factor XI (ASH abstract 12999; addendum: PR on phase I data reported on December 12) and Apo C III programs, there is little doubt left that phoshorothioate-based RNaseH antisense as developed by this company and Santaris can mediate target-specific gene knockdown in the liver in Man. These results confirm the clinical experience with the registrational hypercholesterolemia candidate mipomersen and are corroborated by the impressive HCV results obtained by Santaris’ with its anti-miR122 HCV candidate. Beyond the liver, RNaseH efficacy has been demonstrated for solid cancer (custirsen) and possibly Excaliard’s (now Pfizer’s) anti-scarring candidate. On the other hand, the recent trial termination(s) by Santaris, and the safety profiles of mipomersen and OncogeneX' custirsen highlight some of the challenges facing phosphorothioate antisense technology.

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Therefore, as RNAi Therapeutics have progressed from the basic discovery of its mechanism in mammals 10 years ago to solid proof-of-concept gene knockdown in the form of the ALN-TTR01 results 2 weeks ago, it may be a good time to compare and contrast these two technologies also in light of the fact that, after the RNAi Therapeutics backlash, there is a clear trend towards Big Pharma (and other pharmaceutical companies) opening themselves up again towards antisense, meaning that the 2 technologies are competing for precious non-dilutive funding. For this purpose, I will focus on the liver as the best developed target organ for both these technologies.

20 to 100-fold more antisense required

An obvious advantage of antisense, with about 3x the age of RNAi Therapeutics, is that more is known about its clinical pharmacology. As such, there is good visibility as to how much antisense will be needed to achieve the kind of 50-75% knockdown that will be required for therapeutic outcomes in most cases. Dose has important implications particularly in terms of safety and cost.

Current ‘2nd gen’ molecules (fully phosphorothioated gapmers) like mipomersen require 200mg oligonucleotide per week. Actually, if 300-400mg would have been better tolerated, the higher dosages would have enhanced the commercial profile of mipomersen considerably. Let’s therefore say 1000mg per month for 2nd gen RNaseH antisense.

With the higher-affinity ‘2.5 gen’ technologies that are starting to move into the clinic, best exemplified by Santaris’ LNAs which in fact may also symbolize the most potent version, it is expected that clinical dosages can be further reduced. Based on the non-human primate data and clinical dosage regimes, I expect that dosages of around 100mg/week or 500mg per month are feasible in the foreseeable future. Also because of the modifications involved, I would be therefore very surprised if the cost of oligonucleotides for treating a patient over a year would be below $15,000 even at commercial scale.

By contrast, it can be expected that it will take about 0.15mg/kg of siRNA formulated in the ‘2nd gen’ SNALPs that are now moving into the clinic to achieve the type of once-a-month pharmacology that Tekmira and its licensees are aiming for. If you do the math, that translates into about 10mg per month of siRNA oligonucleotides. Give and take the added costs of the lipids and formulation process, but cheaper nucleotide chemistries involved, this translates into a maybe 50-fold cost of goods difference alone. For some diseases and in some countries, this may be less of an issue, but it will be a factor for others.


The even larger implications of dosage is the related safety. Although clinical repeat-administration studies with SNALP have yet to be conducted, it seems that with the 2nd gen SNALP formulations, the main safety challenge with SNALP will be in managing acute hypersensitivity reactions around the time of drug administration. Based on similar issues with intravenously administered biologics such as monoclonal antibodies where e.g. transient immune suppression with steroids is routine (and widely accepted), I believe that infusion-related acute toxicities will be manageable.

What is nice with the pharmacology of SNALP RNAi Therapeutics is that the bulk of the drug that does not hit the target, i.e. gets incorporated into the RISC silencing complex, is rapidly turned over by the body, meaning that drug exposure levels between drug administrations will be extremely low. It is because of this that I am hopeful that the risk of causing liver toxicity, long believed to be the main toxicity challenge for SNALP, is quite limited at dosages of 0.15mg/kg/month.

By contrast, RNaseH antisense do not harness a natural gene silencing mechanism and, in the case of the phosphorothioate-based gen 2 and gen 2.5 antisense, work by saturating the target (and off-target) organs with high levels of the ‘sticky’ phosphorothioated oligonucleotides so that mass action carries enough of them into the cells. Consequently, the exposure of the body to the antisense drug is significantly higher compared to SNALP-delivered siRNAs. Assuming ~100-300mg of antisense oligonucleotide per kg of liver or kidney tissue (e.g. ISIS TTR patent application US 2011/0294868) and single-digit microgram siRNA oligonucleotide per kg of liver tissue (e.g. Landesman et al. 2010), you could argue that the real difference in bioburden between antisense and SNALP RNAi is about ten thousand fold. It also does not take into account that it is the sticky phosphorothioate chemistry that is thought to be responsible for much of the toxicity (interactions and turnover) whereas RNAi triggers employ more natural chemistries. On the other hand, double-strand RNAs are recognized by more innate immune receptors than highly modified, single-stranded oligonucleotides.

Route of administration

Although the subcutaneous administration of SNALPs has been demonstrated (see e.g. Tekmira's ApoB patent) and may become practical with the higher potencies of SNALPs and extracellular matrix-degrading technologies as developed e.g. by Halozyme, antisense is currently more amenable to subcutaneous administration whereas SNALP have to be infused in an institutional setting. This means that, as is the case for essentially all monoclonal antibody drugs, SNALP drugs have to address diseases of considerable unmet medical needs where patients do not perceive a once-a-month trip to the infusion center a huge burden. Maybe Pfizer can't, but I can think of many such diseases. Infusion in an institutional setting also has the advantage that acute toxicities, the main safety challenge for SNALPs, can be well managed through professional supervision, whereas patients that inject themselves with antisense at home may be slightly panicked on seeing redness develop at the injection site or on experiencing ‘flu-like symptoms’ that have been reported to occur at high frequency with antisense (often 1/3 to 1/2 of patients), but has surprisingly been little discussed by ISIS Pharmaceuticals.


Like route of administration, manufacturing is considered to be a practical advantage of antisense over SNALP RNAi. I agree…in purely practical terms. What is, however, entirely forgotten is that as long as you can deal with the manufacturing complexities, it suddenly gives you an invaluable competitive advantage. How about unlimited market exclusivity? Isn’t one of the lessons that Big Pharma should have learned from the current patent cliff that simple small molecule chemistries are highly vulnerable to generic competition? Isn't this also a major reason for why everybody obsesses about monoclonal antibodies these days, yet is often strangely held against SNALP RNAi? To my knowledge, there are no generics of a nanoparticle-formulated drug.

So in summary, as antisense has reached an inflection point as a slew of clinical data is confirming the early clinical results with mipomersen from 6-7 years ago which demonstrated gene knockdown in the liver, SNALP RNAi is making even faster progress with many of its theorized advantages, especially related to the amount of oligonucleotide required and pharmacology, turning into clinical reality quickly. The race is on. The most likely winners meanwhile are the patients.

By Dirk Haussecker. All rights reserved.

Disclaimer: This blog is not intended for distribution to or use by any person or entity who is a citizen or resident of, or located in any locality, state, country or other jurisdiction where such distribution, publication, availability or use would be contrary to law or regulation or which would subject the author or any of his collaborators and contributors to any registration or licensing requirement within such jurisdiction. This blog expresses only my opinions, they may be flawed and are for entertainment purposes only. Opinions expressed are a direct result of information which may or may not be accurate, and I do not assume any responsibility for material errors or to provide updates should circumstances change. Opinions expressed in this blog may have been disseminated before to others. This blog should not be taken as investment, legal or tax advice. The investments referred to herein may not be suitable for you. Investments particularly in the field of RNAi Therapeutics and biotechnology carry a high risk of total loss. You, the reader must make your own investment decisions in consultation with your professional advisors in light of your specific circumstances. I reserve the right to buy, sell, or short any security including those that may or may not be discussed on my blog.