What TKM-Ebola Could Achieve in the Current Outbreak

(please read financial conflict at the end)

We are in the midst of the largest recorded outbreak of Ebola hemorrhagic fever virus ever and there is little evidence that its spread is being contained.   Here, I will make the case how TKM-Ebola, the most advanced Ebola therapeutic in clinical development, could help in avoiding ever more damage from the deadly virus.

How TKM-Ebola could help

1.       Give suspected cases an incentive to go to the treatment centers.

2.       Provide medical personnel with a stand-by and therefore help in their recruitment.

The continued spread of Ebola can be partly attributed to a breakdown in confidence in the authorities and fear by medical personnel. 

I have been viciously attacking the World Health Organization (WHO) that by downplaying the significance of the outbreak it has been a key factor in re-igniting viral spread.  It is the WHO who have tried to minimize the true numbers of infected, possibly in cahoots with local authorities, and laughed off the suggestion that the virus could get on a plane in arguing against any kind of travel restrictions.  Of course, it recently did and I am still waiting to see top WHO officials send their families on a vacation to Western Africa.  But probably the most outrageous insult was in suggesting that the spread of the virus is explained by ‘funny’ cultural practices in these countries such as kissing the dead during burials.

The WHO, probably in the comfort of their headquarters in Geneva, even attacked on-the-ground Medecins Sans Frontieres as alarmist for calling the outbreak ‘unprecedented’ in late March.

This notion that the virus is very bad in spreading from person-to-person and can only do so with the help of obscure practices is obviously wrong given that more than 100 medical personnel have become infected. While I am still waiting for an explanation by the WHO of how this could happen, I refuse to believe that they kissed the dead in the treatment centers or licked any other of their body fluids for that matter.  Has the WHO (and others) maybe considered the unthinkable, namely that the reason why this is the biggest ever spread of the virus is because the virus has mutated and new routes of infections are possible, such as by aerosol?  The US military will have its reason to believe that this could happen, otherwise why would they be so concerned about it being weaponized and spending hundreds of million dollars on the development and stockpile of an Ebola therapeutic such as TKM-Ebola?

And if people that protect themselves with space-suits get infected, how would you feel as a suspected case of Ebola? 

I know I would do anything NOT to go to these treatment centers, because what is obviously for the good of the overall population would only exponentially increase my risk of contracting the infection in case I was one of those wrongly suspected to have Ebola.

In my opinion, providing individual isolation wards with the best medical equipment possible, a dignified environment, and a drug as an option for the patient could make the difference in whether suspected cases will turn themselves in or not.  The argument that ‘this is Africa’ and you cannot expect good medical care there should not count in this day and age when equipment can easily be shipped between continents.  It’s probably far cheaper to do it now than further risking for the virus to go global (obviously, the WHO thinks this is impossible).

And for medical personnel, the benefit of making TKM-Ebola available is obvious and most tangible: since the onset of flu-like symptoms in this population is highly likely to be due to Ebola and because they have ready access to the necessary equipment such as infusion apparatus, they could be treated immediately with the agent.  Treating as soon as possible is thought to be critical for TKM-Ebola to be efficacious.

What is TKM-Ebola?

TKM-Ebola is an intravenously infused RNAi Therapeutic that has been demonstrated to save the lives of monkeys infected with an otherwise fatal dose of Ebola.  It is being developed under the ‘Animal Rule’ in efforts funded by the US government which is afraid that this virus could be weaponized and used as a bioterror agent.  The ‘Animal Rule’ is a development pathway instituted by the US FDA for diseases such as Ebola for which it would either be impractical or unethical to conduct efficacy studies in humans. 

Because natural outbreaks are unpredictable and experimentally infecting volunteers with the virus out of the question, these monkey studies are as good as it gets regarding drug efficacy (so much for the mantra that there are ‘no drugs for Ebola’).

An important second element of the Animal Rule is that human volunteer studies demonstrate acceptable safety at the doses corresponding to the efficacious dose in monkeys.  For this reason, pivotal phase I safety studies have begun this year in healthy volunteers (so much for the notion that licensed Ebola drugs are far off- ‘phase I’ is misleading).

Clinical Hold

Unfortunately, in the midst of the outbreak, the FDA instituted a Clinical Hold on the TKM-Ebola safety study because a case of dangerously high cytokine elevations was observed at the highest dose planned in this dose-escalating/dose-finding study (0.5mg/kg).  I agree that this is to be considered a serious adverse event in a volunteer that is not infected with the virus.

The reason for the cytokine stimulation is likely due to TLR-mediated, lipid-amplified innate immune stimulation, a known risk of liposomal RNAi delivery, especially at doses of 0.5mg/kg and higher.  It is also the reason why all other active development candidates by Tekmira and their licensee Alnylam are conducted in the presence of transient immune suppression with steroids and the like which in many cases is acceptable given the severe diseases these treatments go after.

As indicated, the adverse event at 0.5mg/kg should by no means spell the end of TKM-Ebola.  Firstly, the company argues that the pharmacologic corresponding dose to those curing the monkeys is lower than 0.5mg/kg.  Secondly, a side effect that is not tolerable in healthy volunteers (usually ~20-year old male students) could be well tolerated in subjects with a 70-90% likelihood of dying from a disease in a matter of days.  It is ethically more troubling to involve healthy volunteers in such drug development just as aggressive experimental cancer drugs are hardly ever tested in healthy volunteers.  Lastly, there may be ways to avoid the side effect altogether, such as by using transient immune suppression.  However, I do not know whether transient immune suppression is possible for Ebola, but I expect Tekmira will have the answer for this from their large-scale animal experience.

Next steps

For TKM-Ebola to have the best impact on the current epidemic, the first step would be to start manufacturing it at scales sufficient to treat at least ~1000-5000 patients.  This takes time and given the uncertainty around the future course of the epidemic, the investment needs to be made now instead of waiting until it is too late for a treatment center-focused approached involving an intravenously infused agent.

The next step depends on the feasibility of steroid pre-treatment during an Ebola infection.

In case that it is known that steroid pre-treatment was of little concern (e.g. based on infected monkey studies), start treating rather aggressively (e.g. start at doses of 0.25mg/kg).  In case it was not, go about more slowly by starting at sub-therapeutic doses as low as 0.025mg/kg and treat the initial experience like a dose-escalation study in actually infected patients.  Of course, everybody would need to provide informed consent.  In general, any semblance that the use of TKM-Ebola was imposed by the Western world and the local population used as guinea pigs is to be avoided which is probably a key reason why Tekmira to my mind has been almost in hiding during this whole episode almost to the point that they (and the FDA) are glad about the Clinical Hold.  

To support the accelerated development of Ebola therapeutics and vaccines, including TKM-EBOLA you can add your signature to the following change.org initiative: http://www.change.org/en-GB/petitions/food-and-drug-adminstration-fast-track-drug-and-vaccine-research-for-ebola-hemorrhagic-fever

Disclosure: Tekmira constitutes a meaningful part of my investment portfolio and I have to credit the WHO for greatly increasing its value.

Quark’s Failure in Kidney Study Heralds End of Naked RNAi Triggers

Quark Pharmaceuticals today announced that RNAi Therapeutic candidate QPI-1002 has failed in a fairly large phase II kidney transplant study to meet its primary efficacy end-point.   For those interested, it was a pre-defined threshold of reducing the risk of having to go on dialysis after receiving a kidney transplant (30% pre-defined, 15% achieved with no p-value provided).  Following failures with similar 'naked' RNAi candidates by Quark and others (e.g. Opko/Acuity wet AMD candidate and Alnylam’s ALN-RSV01), this probably puts a nail in the coffin of a naïve, but convenient approach to RNAi delivery taken by quite a few in the early days of RNAi Therapeutics. 

QPI-1002 is a blunt-end 19 base-pair AtuRNAi targeting p53, thereby aiming to protect struggling kidney cells trying to take hold in their new host from dying.   It carries simple alternate 2’-o-methyls on both strands and is given by intravenous infusion reconstituted in saline.  While it is well established that unformulated/naked oligonucleotides without special modifications or delivery formulations to make them ‘drug-like’ will concentrate in proximal tubule cells of the kidney, the declared target cell for QPI-1002, it is still a mystery how the RNAi trigger is supposed to cross into the cytoplasm of those cells. 

Of course, a drug may fail for reasons other than target engagement, which couldn't be determined in this study, but I believe Quark in private would also side with the view that this most likely had to do a failure to deliver. This is because Quark itself has been adding chemistry to their more recent molecules to turn them into more credible self-delivering RNAi triggers.  And even if it wasn’t due to a failure to deliver, there is now little reason to pursue it any more.

RIP naked RNAi, RNAi Therapeutics has moved on.

Dear Chris Garabedian,

I am writing to you today to introduce myself as a potential candidate for the CSO position at Sarepta.  Based on media reports last week (see here for the Wall Street Journal version), it seems that this position has become vacant after you had a little spat with Art Krieg.  If the anonymous sources close to Sarepta cited so abundantly in those reports were to be believed, at least some of the disagreement related to Art’s failure to push and expand the morpholino-based product pipeline the way you would have liked.  Art maybe be an icon in the oligo industry, but I believe that I am the right man for you as I know where the gold mine for morpholino oligonucleotides lies.

First, however, a few words about myself.  You may have read or at least heard about my critical writings of your company’s pursuit of getting eteplirsen, your exon-skipping drug candidate for Duchenne Muscular Dystrophy, on the market.  In case that there was some truth to speculations that some of the disagreement related to the accelerated approval strategy for eteplirsen, rest assured that I will ignore all my concerns about the conduct of the rather small trial and biochemical assays used, and tow the party line anyway.  Even better, I would be prepared to cede any involvement in the experimental conduct and interpretation of data generated in preparation of filing for accelerated approval although these activities would naturally call for my involvement as the CSO of the company.

Coming back to my prior criticisms of the company….to be frank it was Janet Yellen and the adverse impact her comments have had on biotech stocks that now forces me to apply for employment.  Nothing personal, no hard feelings, we will all be able to get along and laugh about it later.  I am running out of options as I have burned down bridges with virtually all other RNA Therapeutics companies in the field as I have freely criticized them before when I still had money.  As you know from reading the message boards carefully, I have been desperately applying for jobs there only to be rejected and attack them even more venomously.  Given my track record and financial situation, regardless of the fates of both Krieg and Linsley before that, please forgive my youthful follies and I fully expect you to be a difficult and demanding boss and I will put up with it.

I know that you have long made up your mind to not hire me and the only reason for humoring me and reading on is the thing about the gold mine.  I genuinely believe there is one and- you are right- it’s got to do with splice modulation and by extension the modulation of RNA processing by masking sites for RNA binding proteins.  In particular, I find morpholinos highly attractive for splice modulation in pharmacologically contained compartments such as the CNS where the risk- and please excuse my language here- that the expensive oligo is pissed out immediately is limited.  There is a reason why morpholino has become such a popular tool to study developmental biology in zebrafish embryos and the like: they cannot pee!!!  So with the ability of maintaining high tissue concentrations and the amazingly safe morpholino chemistry, you may be able to generate invaluable medicines for awful diseases such as spinal muscular atrophy (maybe even better/safer than what ISIS is generating currently) and Huntington’s disease.

Kind regards,

Dirk. 

Fed-Induced Sell-Off Offers Attractive Values in RNA Therapeutics

For whatever reason, Janet Yellen does not like cheerful biotech investors and between her first policy speech as new chairwoman of the Fed in April and comments last week decrying biotech and social media stocks as inflated she caused two major sell-offs in RNA Therapeutics stocks.  As a result, industry bellwethers ISIS Pharmaceuticals and Alnylam are trading at less than half their 52-week highs, though better than ‘second-tier’ RNA Therapeutics companies such as Tekmira and Arrowhead Research which are trading down even more than that.

When the technology has matured to the point that it can bring significant value to patients, pipelines rapidly progress and expand, and with companies flush in cash by historical standards, this sell-off offers in my opinion great risk:rewards.  Here is a run-down of my five favorite plays.

1.       $ISIS. ISIS Pharmaceuticals is my current RNA Therapeutics core holding that I would not want to be out of.  With ligand-targeted technologies offering much lower tissue exposures and therefore much better safety margins, multiple mechanisms of actions and a pipeline that rivals any in the industry, this company is on track to become the most valuable in the industry- if not economy (subject to the vagaries of the healthcare policy directions). 

At a $3B market cap, no foreseeable financing needs, and a healthy newsflow (especially regarding partnerships), the downside should be relatively limited, and I will sleep tight even knowing that Yellen (who seems to confuse the Dow and large cap biotechs with the broad biotech industry) might open her mouth again to talk about something she obviously understands very little about.

Best long-term risk/reward.

2.       $TKMR. Tekmira is the most beaten-up stock of the bunch that gets credit for almost nothing.  This is mainly the result of the Ebola trial results which were a disappointment- little doubt about that.  However, translating these results from the unthankful ‘Haertetest’ to the rest of the pipeline is unwarranted.  This is because a highly most potent knockdown technology for the liver, should be able to carve out some market for itself.  And if the HBV product won’t require transient immune suppression and/or if its use is deemed acceptable in HBV with its expected finite treatment period, there is an enormous opportunity for which the favorable newsflow is only about to heat up with the presentation of the preclinical dataset..

It is the newsflow from new product candidates (esp. HBV)), clinical trial starts (liver cancer, HBV, alcohol dependency), clinical trial results (ALN-TTR02 OLE, TKM-PLK1 interim phase II with little to no expectations by the markets), and business development (mRNA), that makes TKMR an attractive buy at these severely depressed levels ($31 in April, now trading at $9).  And with the Ebola Haertetest on Clinical Hold, TKM-EBOLA does not scare me anymore as an investor.  It even offers a potential near-term catalyst should the Hold be lifted.

Best 6-12 month risk/reward (until release of phase IIa HBV knockdown results).

3.       $RGLS.  I’ve never liked Regulus much as a stock, mainly because of its anemic pipeline that its own management does not even seem to be excited about given that it now calls the potent GalNAc-targeted anti-miR122 product candidate for HCV merely a proof-of-concept for the company.  However, given that the HCV market emerges as one with a long-tail, not all is lost for this pan-genotypic, potentially one or two-shot treatment when used in conjunction with other HCV drugs. 

When HCV-competitor Achillion recently surged on the back of the Merck takeout of HCV play Idenix as it reminded investors of the long-tail nature of the market, Regulus was forgotten.  It may reach investors’ consciousness when they report phase I results possibly early next year.  Solid financials with more than $100M in cash and a reasonable $300M market cap given its prominent position in the microRNA Therapeutics industry.

Speculative buy with regard to single catalyst.

4.       $ARWR. All eyes are on the outcome of the ARC520 phase IIa study in HBV-infected patients which could be announced any day now.  My prediction is a ~80% HBsAg knockdown at the 2mg/kg dose.  My problem is that I have little idea about how the market will respond to that given that the trial is not designed to achieve the ultimate goal for this therapeutic candidate: a functional cure.

Most investors seem to expect an easy double (or more) from current levels, an notion that I do not want to necessarily reject for an 80% knockdown.  It all seems to depend on how the results will be framed by the analysts and how successful the dueling hedge funds are in painting the tape to their advantage. 

With more than 10% of the float short and given ARWR's historical trading pattern, great volatility is a given.

Highly speculative buy, so much so that options may actually be the best way to play it.

5.       $SRPT. No matter the negative market reception to the recent release of the 144-week study results, Sarepta and patients will get their accelerated approval for eteplirsen.  The debate has reached a point that the drug development clock for Duchenne Muscular Dystrophy cannot be set back another 5-10 years by rejecting the current class of candidates of which Sarepta’s looks to be the most promising- despite the highly anecdotal nature of the results.  This is because eteplirsen seems amazingly safe, so the potential harm from giving a potentially non-effective medicine is limited.  This is clearly not the case for Prosensa’s exon skipping drug candidate, and don’t even get me started about PTC Therapeutics …

Solid 12-month risk:reward juiced by the large short position (>30%).

GalNAc Advance by ISIS Means Oral Oligonucleotide Therapeutics around the Corner

Last week, the long-awaited first publication on ISIS’ GalNAc-targeted antisense oligonucleotides appeared in a ‘NAR Breakthrough Article’ (Prakash et al., 2014)  Living up to that label, the data with RNaseH antisense oligonucleotides (ASOs) conjugated to triantennary GalNAc sugars showed that this strategy increases potency by nearly an order of magnitude.  In combination with high-affinity chemistries (in this case cET) that facilitate the use of short (12-14nt vs ~20nt) oligonucleotides which has been shown to improve in vivo potency (Santaris research by Straarup et al., 2010), oral delivery for antisense modulation of hepatic gene expression has essentially been solved. 

Moreover, the lower dosages that can now be used in addition to the improved biodistribution profile that is now heavily slanted away from non-hepatocytic cell types in the liver and the kidney in favor of hepatocytes, means that the other major benefit of GalNAc-ASOs with immediate applicability is that the safety of therapeutic ASOs for liver-directed applications will be much improved.

The conclusion that oral antisense therapeutics are just around the corner is based on the observation that a relatively crude caprate-based phosphorothioate oligonucleotide formulation enabled a ~10% bioavailability following oral administration already (intravenous = 100%; Tillman et al., 2008).  This, in addition to some inter-subject variability that might have been linked to gastric emptying times, meant that too much oligonucleotide would have had to be administered orally with previous antisense oligonucleotides. 

Granted, this first piece of GalNAc-ASO literature did not explore oral delivery. The fact, however, that the combined use of GalNAc conjugation with high-affinity chemistry improved parenteral ASO potency by 60-fold is predicted to mean that oral GalNAc-cETs are already more potent (~6x) than subQ-administered 2^nd gen ASOs.  These have already produced impressive phase II data for targets such as Factor XI and ApoCIII.

The potency improvement in terms of oral delivery may actually be larger than 60-fold.  This is because oral delivery strategies such as caprate co-formulation which aim at increasing intercellular drug permeability are size dependent.  This means that there should be an added benefit of the smaller oligonucleotide size facilitated by cET chemistry with oral delivery.

Of course, the data leave open some questions.  My most pressing, applicable to both oral and non-oral uses of GalNAc-ASOs is how the rodent data translate into non-human primates and ultimately humans.  This is because for unconjugated phosphorothioate oligonucleotides, the potency on a mg/kg basis improves with the size of the organism.  The discussion above is based on an assumption that GalNAc-targeting will not change that.  I have been unable to make a determination yet as to whether this should hold true or not.

Other than that, the antisense weather forecast is blue skies ahead, though with a chance of thunderstorms in the form of IP skirmishes with their good friends over at Alnylam as they have pioneered GalNAc conjugation in oligonucleotide therapeutics, but may now be ironically penalized for it from a competitive point of view.  Alnylam is not the type of company that just lets competitive threats happen to them.

Implications of Ebola data for Tekmira HBV Program

Like it or not, it is the HBV program that is driving the interest in Tekmira, at least in the near-term.  Therefore, the 15% sell-off in the stock yesterday following the Clinical Hold imposed by the FDA can be interpreted to be due to the uncertainty of what it means for the HBV program.

It is my working hypothesis that for the treatment of HBV via the immune reactivation pathway, the use of even transient immune suppression is to be avoided.  On the other hand, the recent Ebola clinical results by Tekmira suggest that transient immune suppression may also be desirable for 3^rd gen SNALP delivery.  As TKM-HBV is expected to utilize '3^rd gen' SNALP delivery, too, the Ebola results with immune activation seem like a bad omen.

While this cannot be ignored, there are a few reasons to believe that this might be too simplistic.

1.       Difference in dose

Although there was an apparent tendency for immune stimulation throughout the dose escalation with TKM-EBOLA, it was only at the high dose of 0.5mg/kg where a dose-limiting toxicity was observed. And according the yesterday's press release on the Clinical Hold, it is in particular cytokine elevations that were observed at 'higher doses' that are the issue.  

 Since unlike Ebola, HBV only replicates in hepatocytes, and because SNALP potency is highest in hepatocytes, the efficacious dose for HBV may be well below 0.5mg/kg with 3^rd gen formulations.  Of note, even 2nd gen SNALPs are highly effective at 0.3mg/kg based on the Tekmira-enabled ALN-TTR02 candidate now in phase III.

2.       Different sequence, different immune stimulation

It is known (e.g. Judge et al. 2005: Sequence-dependent immune stimulation of the mammalian innate immune response by synthetic siRNAs) that immune stimulation by RNAi, especially when liposomally formulated, is highly dependent on the actual sequence and modification pattern of the RNAi trigger.  Since TKM-EBOLA and TKM-HBV are different in that regard, it is well possible that while one 3^rd gen formulation is immunostimulatory, the other is not.

3.       Immunostimulatory by design/by screen

Scientifically, I was most disappointed by the Ebola immune stimulations as Tekmira made a big deal in the wake of similar observation in 2010 with TKM-ApoB that with improved, more predictive immune stimulation assays, they would now be able to screen out the immunostimulatory formulations.  In a black-and-white world you would conclude ‘obviously not!’.

In a world of shades of grey, I can imagine that TKM-EBOLA might have happened to slip through this part of the screening process.  The reason is that since Ebola infects a number of cell types besides hepatocytes (endothelial cells, phagocytes etc), the impressive efficacy of TKM-EBOLA always stood out as something quite unusual.  So while containing the spread of the virus by hitting it particularly hard in the liver might be part of the mechanism, I am quite receptive to the notion that the scientists did not mind all that much some immune stimulation when the RNAi formulation had so much efficacy.

With HBV, however, the company might be much more careful in screening out immunostimulatory potential.

I can now hear Sarepta investors yell ‘it’s an artefact’, but I would caution them that Sarepta and the morpholino field at large is proposing morpholinos with cationic appendices for infectious diseases while these are not really considered for genetic diseases.  What I don’t understand is that why a modification that is supposed to be helpful in e.g. suppressing viral gene expression should not likewise be useful for suppressing human gene expression.  So maybe the value of these modifications is in tickling the immune system?

4.       Not all 3^rd gen are created equal

In response to a questioner in the Q1 financial conference call, the CEO of Tekmira said that the definitions of the various SNALP generations is not as clean as you would like from a chemical point-of-view.  Instead, the definition largely is a reflection of the potency improvements of the formulations.  Therefore, since TKM-EBOLA has a different set of target cells than TKM-HBV, it is well possible, even expected, that the formulations will also differ in their lipid compositions.  Since lipids are an amplifier of the liposomal-RNAi immune response, differences therein could have big implications as to the immune stimulation of the various formulations.

Disclosure: I am long Tekmira and only partially heeded my own advice of stepping out of the TKM-Ebola ‘Haertetest’ (trial provided not much upside to investors).  On the other hand, Tekmira has upside potential from a number of non-Ebola programs especially HBV and mRNA delivery.  As the worst case scenario, an outright termination of TKM-EBOLA, is a real risk, it becomes more and more difficult to get the timing right ahead of the upcoming TKM-HBV data.  If, however, you don’t mind volatility and are a long-term investor, it’s another matter.  However, at least for me, Tekmira is not the stock any more to maintain a 100% position in.

Tekmira Ebola Candidate Halted as Disease Spreads in Western Africa

There could not have been a worse timing for the Clinical Halt imposed on Tekmira’s RNAi candidate for the treatment of Ebola viral infection, TKM-EBOLA.  Just as the disease is reaching epidemic proportions in Western Africa (>500 dead), the world’s leading drug candidate for this infection with validation in the most stringent monkey models got stopped in its tracks following concerns by the US FDA regarding its immunostimulatory potential that was evidenced in the single-ascending dose part of the study (discussed here).

Efficacy in monkeys is the best you can hope for in the absence of an actual outbreak such as the current one, so I was hopeful that TKM-EBOLA could become part of the solution, especially by providing an incentive for patients to identify themselves and come to the treatment centers.  It is the prospect of dying surrounded by people in space suits instead of loved ones, and even worse the prospect of being infected by Ebola in the first place from fellow ‘inmates’ if you have been misdiagnosed that keeps people away from the hospitals.  This appears to have been key to the continued spread in addition to cultural practices such as kissing the dead as the WHO likes to emphasize (in my opinion, the condescending attitude by the behind-the-curve, out-of-touch WHO has been the 3rd key factor).

I suggest to mobilize the financial resources to fly in the latest medical equipment and offer patients to be treated with something that might help them survive.

Instead, immune-related side effects such as flu-like symptoms in healthy volunteers made the FDA ask Tekmira to provide additional data and explanations to justify proceeding to the multi-dose ascending part of the study conducted under the Animal Rule regulatory pathway.

In a way, this development might be considered support for those that have said that it is too early to treat patients with TKM-EBOLA, or any other experimental treatment for that matter.  On the other hand, you could also argue that this only shows that it is unethical to test it in human volunteers where the tolerance for side effects is much less than in somebody with a ~70% risk of dying from an infection otherwise.  Just look at the cancer drug development field, and at least in RNAi Therapeutics, I have never seen them tested in healthy volunteers even in phase I, but straight in cancer patients that have little to lose.

In fact, the immune stimulation could be the price to pay for a cure as it might be part of the mechanism of action of TKM-EBOLA either by shutting down viral protein translation and/or by interfering with the virally-induced cytokine storm itself.  These scientific issues, also as it relates to Tekmira’s platform in general, will be discussed in a follow-up post.

Test of Tekmira's credibility

I would like to close with some remarks on the credibility of Tekmira’s management.  

Management did in fact disclose the risk that there might be delays with the phase I study following results from the single-dose part of the study when it said there would be FDA review ahead of the multi-dose part of the study: 'Interim safety data submitted to the FDA for review May 7, 2014.'

Although this could be read as part of the normal regulatory process, the fact that they disclosed it made me believe that this was a real possibility.  The positive spin on the data obviously falls under the Safe Harbor of ‘forward-looking statements’ and no foul play here.  In fact, many biotechs would not even have disclosed the FDA review. 

On the other hand, many investors went along with the positive sentiments by management and will feel disappointed now.  In order to redeem themselves to this constituency, management will now have to prove that they were already prepared for this event and that they can speedily answer the questions and provide the requested data so that the trial may resume asap and that TKM-EBOLA may be offered to those in Western Africa now that it can be considered that the drug has been fully vetted for safety.

RNAi Trigger History Revisited as Alnylam Once Again Gets Tough on IP

Last month, Alnylam lectured  Arrowhead Research that they infringed Alnylam's RNAi trigger IP and added that the use of UNAs would not protect companies using them (Tekmira, Arrowhead Research, Marina Biotech, and Arcturus) either.  So as Alnylam is once again trying to intimidate the competition and their investors by claiming that they control RNAi triggers, it might be worth revisiting some RNAi trigger history.  Doing so suggests that in fact they have been one of the egregious infringers of 3^rd party IP themselves and raises questions as to their own freedom-to-operate and validity of IP.

Sirna and Silence Therapeutics early proponents of modified RNAi triggers

The early RNAi days were marked by the competition for RNAi trigger supremacy between star-studded start-up Alnylam and working-class Sirna Therapeutics which had just averted bankruptcy following failed attempts with ribozymes.  Silence Therapeutics was another company that had ambitions to cornering important RNAi trigger IP and like Sirna had a history in failed attempts with oligonucleotide therapeutics.

Due to their prior experiences in nucleic acid-based drug development, it came natural to Sirna and Silence to apply nucleic acid modifications and soon earned a head-start in creating IP related to the use of modified RNAi triggers.  The belief was that in order to endow RNAi triggers with pharmaceutical properties, they had to be modified.

Sounds familiar? If so, then it is probably because this has become Alnylam’s new mantra, too.  Of course, this comes with a 12-year delay and only after acquiring Sirna’s IP estate and a history of claiming that they want their RNAi triggers to be as ‘natural’, i.e. unmodified, as possible, not due to IP limitations, but because of superior performance and considering safety.  In fact, Alnylam has not only been infringing Baulcombe IP for a short while, but also Merck/Sirna IP for probably as long as they were starting to see decent in vivo knockdown results with their highly modified GalNAc-siRNAs.

Although this clearly shows that Alnylam’s publicly claimed control over RNAi trigger IP has simply been a mirage, to their credit they make efforts in eventually gaining access to IP they are glaringly lacking.  Also, while at some point and for some delivery strategies the notion of minimally modified RNAi triggers had its merits, the emergence of RNAi trigger conjugates has increased the value of RNAi trigger modifications.  One might also criticize companies like Sirna/Merck, Silence, and RXi Pharmaceuticals (one of the first practical adopters of highly modified RNAi triggers and conjugates) for standing still instead of developing their initial technologies to their logical conclusion.

A comeback for AtuRNAi triggers?

Silence Therapeutics left a lot of value on the table by limiting themselves to a modification pattern that involves alternating 2’-O-methyls with a 2’-O-methyl-modified base on one strand opposing an unmodified base on the other like in a zipper.  One reason for this limitation was that during patent prosecution, the attempt by Silence Therapeutics to get patents related to modification patterns in general and not limited to 2’-O-methyls failed.

Interestingly, the RNAi triggers in ARC520 that Alnylam lays claim on involve a modification pattern reminiscent of AtuRNAi, only that it is here 2’-fluoro that is the alternating modification (Wooddell et al. 2013).  Also, if you review Alnylam’s recent patent applications (e.g. WO2014089313), alternating 2’-fluoros seems to be Alnylam’s preferred modification pattern, too.  Could it be that these companies were inspired by AtuRNAi trigger design?   

In any case, AtuRNAi and Silence may hold the key to invalidating some troublesome modified RNAi trigger IP by Alnylam by rendering them obvious.  Even more exciting for Silence, albeit unlikely given that the opportunity may have passed is the prospect that Silence could revive some of the more general RNAi modification pattern claims, thereby affecting the freedom-to-operate by Alnylam (and Arrowhead Research).

And welcome to Amy Shuman, former general counsel of Pfizer, to the Board of Directors of Alnylam.

Leveraging New RNAi Trigger Chemistries for Gene Knockdown in Phagocytic (and Other) Cells

Much of the achievement of solid RNAi gene knockdowns in hepatocytes (liver) by non-LNP means (Arrowhead DPCs and Alnylam’s 2^nd gen GalNAc-siRNAs) has involved the use of heavy modifications that render the RNAi triggers highly stable. This is because nucleic acids that are not protected by the delivery chemistry itself would otherwise be subject to rapid degradation in extracellular body fluids.

In the case of Alnylam’s GalNAcs, they can even function in the absence of an explicit endosomal release chemistry.  Moreover, GalNAcs and DPCs have shown more sustained gene knockdowns than is achieved with LNP delivery which historically has relied on minimally modified RNAi triggers. 

All that is required is receptor-mediated uptake into the endosomal-lysosomal pathway which is an immensely degradative environment (esp. the lysosomes).

Considering the extended duration of silencing and degradative environment, it seems as if the RNAi triggers have to be able to survive for long enough in late endosomes/lysosomes so that when they get an opportunity to escape by as yet undefined mechanism(s), they are still there ready for gene silencing action.

Such chemistry progress may also be particularly useful for RNAi gene silencing in phagocytic cells of the immune system.  This is because there have been multiple reports, especially concerning the use of LNPs (e.g. Novobrantseva et al.)  where some, but not the very robust, hepatocyte-type of gene knockdown have been obtained.  It is the ability to confidently achieve robust knockdowns that opens the gate to a flood of therapeutic applications, and this is why pushing borderline-technologies over the edge is so tremendously valuable.

Uptake into phacocytes is usually not the problem as these have evolved to scavenge for foreign particles and macromolecules.  In fact, phagocytic uptake is often a nuisance in RNAi delivery both because it may cause off-target toxicity and because it can make pharmacology less predictable.

In the case of untargeted nanoparticles such as LNPs, phagocytosis is the likely uptake mechanism.  Similar to endosomal uptake, phagosomal uptake involves the fusion with lysosomal compartments meaning that phagosomal contents are also exposed to a highly hostile environment.  Since the normal endosomal escape chemistries and mechanisms do not appear to be very effective in phagosomes, one strategy besides of possibly tailoring existing mechanisms to the phagosomal environment (e.g. lipid pKas) is to simply use the same ultra-stable RNAi trigger chemistries that are showing promise in the liver.

It is also possible that ligand-targeted conjugate approaches will be useful here, whether they enter the cells via phagocytic mechanisms or not.  In fact, Arrowhead Research (then Mirus Bio) in their seminal publication on DPCs (Rozema et al.) have demonstrated efficient uptake of DPC-conjugates into liver phagocytes (Kupffer cells) by using mannose as the targeting ligand (they haven’t tested and/or shown the corresponding RNAi knockdown though and this may relate to their using much less nucleic acid modification back in 2007). 

Since the mannose receptor is expressed on phagocytes throughout the body and not just in the liver, more extended circulation times promises the application of this conjugate-targeting strategy more generally.

Similar principles may apply to the targeting of other ‘frontier tissues’ for RNAi delivery.  However, given the degradative and differing nature of phagocytosis which means that other release mechanisms are not readily applied to this process, RNAi in phagocytes should particularly benefit from the new RNAi trigger chemistry developments.  I look forward to seeing the results.