A Therapeutic Dose of RNA?

With news of the partnership between GlaxoSmithKline (GSK) and Arrowhead for the development of RNA-interference (RNAi) treatment ARO-HSD, a candidate for treating nonalcoholic steatohepatitis (NASH), it is a good time to revisit the field of RNA therapeutics process and its wide ranging potential of applications in pharmaceutical and biotechnological environments. 

Mechanism of Action

Currently, there are two major categories of RNA therapeutics:

• RNAi, which uses double stranded interfering RNA molecules that have to interact with enzymes such as DICE to initiate a chain of silencing effects
• Antisense Oligonucleotides (ASOs), which use single stranded RNA that binds directly to messenger RNA (mRNA)

Both modalities function by silencing gene expression at the RNA level. They achieve this by interfering with the messenger RNA transcribed from a gene before the mRNA can be translated to a protein. They can also interfere with other forms of RNA which may have functions beyond coding for proteins. This is in contrast to CRISPR gene editing or other gene therapies which interact with genes at the DNA, rather than the RNA, level. 

RNAi is brought about by microRNA (miRNA) and small interfering RNA (siRNA) molecules, which form a natural part of cellular defense mechanisms against viruses and transposable DNA. It can also be introduced to cells via exogenous means – which is why the phenomenon has wide-ranging applications across a host of fields, as it interacts with one of the most fundamental mechanisms of cellular function. Research has shown siRNA to be capable of antiviral treatments, such as by silencing viral genes integrated into host cells (e.g. with Human Papillomavirus) as well as silencing the expression of receptors viruses use to enter host cells (such as with HIV-1). Naturally, it also has a plethora of applications in cancer treatment – whether through disabling carcinogenic gene products, or suppressing immune inhibitors to assist immunotherapies, among others.

Challenges in the use of RNA

Corollary to these wide-ranging applications come steep challenges that must be overcome. Delivering RNA to intended locations is the primary roadblock, which is why it took nearly two decades for the first RNAi therapy to be approved. Firstly, only small molecules of RNA can be utilised – longer-chain RNA nucleotides can trigger interferon responses, which is an antiviral reaction from the immune system. Treatments must also be administered in larger doses due to the low bioavailability of siRNA and its rapid clearance by cells. As siRNA also contains multiple negative charges, it cannot cross cellular membranes in its natural form. 

The solutions to these delivery problems have involved finding alternative ways of transporting the molecules; the two primary ways to achieve this are to either deliver it inside nanoparticle carriers, or to conjugate (combine) the molecules with compounds that can bind to cellular receptors. Several other chemical alterations can also be performed to alter the rate at which the molecules are excreted, degraded by nuclease enzymes, the rate at which they are released from the carriers that deliver them, as well as how they avoid immune recognition. While these function as roadblocks, they also offer flexibility and opportunities for further manipulation in precisely how long the molecule can remain active, among other things. 

Industrial Developments

Beyond these academic findings, RNAi has also seen developments in industry and real-world applications in the past decade. Alnylam Pharmaceuticals is the leading company working in the field, having developed multiple approved therapies. The first FDA-approved siRNA treatment was patisiran, developed to treat familial amyloid polyneuropathy by preventing the production of the abnormal form of the transthyretin protein that causes the disease. Alnylam has also developed lumasiram and givosiran to treat other rare genetic diseases – which have also received FDA approval.

But beyond rare genetic disorders, Alnylam has also developed inclisiran, which is an RNAi-based inhibitor that prevents the translation of PCSK9, a protein which reduces the degradation of low density lipoprotein-cholesterol (LDL-cholesterol). Disabling PCSK9 can lower blood cholesterol levels, which is what inclisiran achieves: it can produce up to a 50% reduction in LDL-cholesterol levels. The real improvement over current treatments such as statins comes from the fact that inclisiran only needs to be administered twice a year, which drastically improves patient adherence. 

Sirnaomics is another company that leads innovation in the RNAi fields, with their primary treatment candidate, STP707. Sirnaomics focuses on altering tumour microenvironments through the use of siRNA, thereby enabling the immune system to achieve better infiltration of the tumour. STP707 is a combination of two siRNA elements that each target TGF-β1 and COX-2, delivered via nanoparticles. TGF-β1 and COX-2 can promote tumour inflammation and fibrosis, which makes it very difficult for the immune system to interact with the tumour and often leads to the cancer being refractory to standard treatment. Pre-clinical models have shown that STP707 can effectively prevent translation of TGF-β1 and COX-2, and achieve greater T-cell numbers in the tumour environment.

Additionally, siRNA-based therapies administered directly to the bloodstream tend to accumulate in the liver. While this was originally a setback for development strategies aimed at other organs, it has given a boost to therapies targeting the liver specifically. This is the strategy which underpins Arrowhead and GSK’s approach to treatment candidate ARO-HSD. ARO-HSD downregulates the translation of the HSD17B13 enzyme – which is involved in the metabolism of hormones, fatty and bile acids. Genetic data from humans indicates that non-functional HSD17B13 can contribute to significant risk reduction for a plethora of liver conditions – such as cirrhosis, alcoholic hepatitis and non-alcoholic fatty liver disease. 

RNA interference holds promise for areas beyond pharmacology, however. In particular, RNAi-based insecticides have already been engineered and are expected to be important in developing more specific ways to tackle agricultural pests while reducing other ecological and environmental impacts from the crop protection industry. In 2018, Monsanto received approval for a transgenic corn that can express RNAi traits lethal to rootworms. Work is also being carried out in topically applied RNAi-based pesticides that do not require genetic modification of the crop plants, although such products are harder and more expensive to engineer. Similarly, crops can be modified through RNAi to develop other traits beyond pest toxicity.

It is obvious that RNA interference technologies hold immense potential for a variety of industries. For the past two decades, the industry has had a rather on and off relationship with the idea. While their discovery was met with much optimism, as evidenced by the awarding of the Nobel prize in Physiology to the RNAi pioneers Andrew Z. Fire & Craig C. Mello, the difficulties concomitant with their practical application soon became apparent. These include extremely challenging modes of delivery and persistence in their target areas and organisms, as well as questions regarding their specificity. However, many companies persevered despite these setbacks – culminating in the approval of multiple products developed by Alnylam, with other companies now following behind or re-entering the field, such as Dicerna, Sirnaomics, Silenseed – with pharmaceutical giants such as GSK, Novartis and Sanofi increasingly giving rise to partnerships with these innovative companies. Given these big moves in the industry, the sun seems to be rising on the horizon for innovations in RNA interference across the entire sphere of the life sciences.

Nick Zoukas, Former Editor, PharmaFEATURES