Bridging the gap

C&I Issue 8, 2013

Existing pharmaceuticals can only tackle 10–20% of known drug targets, leaving at least 80% ‘undruggable’. Many of these undruggable targets involve proteins inside cells, but our inability to reach them may be about to change. In May 2013, Aileron Therapeutics announced the successful completion of Phase 1 clinical trials of its first so-called ‘stapled peptide’ drug – the first in a potentially game-changing new class.

Currently, there are a limited number of peptide drugs on the market, such as the diabetes drug Byetta. These drugs target receptors on cell surfaces. However, by forming a hydrocarbon bridge between amino acids in a peptide chain, researchers can create drugs that reach previously inaccessible disease targets within cells.

The bridge, or ‘staple’, forces the peptide to adopt the α-helical shape found in proteins. It is this structural change that gives stapled peptides their drug-like properties. As well as penetrating cells, stapled peptides stay active in the body longer than other peptide drugs.

Their helical conformation protects them, because peptide-digesting enzymes within the body only recognise and breakdown unravelled peptide chains.

The field of stapled peptides was pioneered in the late 1990s by Gregory Verdine, the late Stanley Korsmeyer and Loren Walensky, at Harvard University, US, and the associated Dana-Faber Cancer Institute. In 2004, the team published a ground-breaking paper that showed a stapled peptide could block the growth of leukaemia cells in mice (Science, 2004, 305, 1466).

Chemical biologist, Verdine explained: ‘We wanted to maximise the likelihood of cell penetration, and so unlike what everyone else had done previously, we put only hydrocarbon atoms in the macrocyclic bridge.’ The hydrocarbon staple is made by incorporating two amino acids with attached alkene groups into the peptide chain. The alkene groups are then connected using the 2005 Nobel-prize winning ruthenium-based Grubbs catalyst. The staples are generally applied across one or two helical turns.

The resulting stapled peptides combine the desirable features of the two main existing drug classes: their ability to lock-in to target proteins gives them the specificity of biological molecules such as antibodies; and their small size (around 50 amino acids) allows them to penetrate cells like small molecule drugs.

The potential for this new type of drug is large in Verdine’s opinion. ‘The operating theatre of small molecules is about 10% of human targets,’ he says. ‘If we take biologicals… that’s 10%...if stapled peptides can reach another 10%, they would have the impact of the entire pharma or biologics industry.’

Aileron Therapeutics was founded in 2005 by a team including Verdine and Walensky. The start-up’s financial backers include the venture arms of four big pharmaceutical companies – an indication of how seriously stapled peptides are being taken. In 2010, Aileron established a collaboration with Swiss pharmaceutical giant Roche, potentially worth $1.1bn. It includes drug development targets in five areas: cancer, viruses, inflammation, metabolism and the central nervous system.

Stapled peptide design almost always starts with the synthesis of an α-helix sequence from a natural protein, but to be an effective drug, the peptide needs to be cell-penetrating. Stapled peptides are thought to enter cells like viruses, via active take-up.

Aileron is confident it can develop stapled peptides that enter cells. Vice president of corporate development, Huw Nash comments: ‘We have a very good understanding of the structure–activity relationship guiding stapled peptides entry into a cell or what keeps it outside the cell…we’ve learnt the rules that help optimise that process.’

Verdine describes optimisation to find the best drug molecule as ‘the heavy lifting part’. ‘You need to screen a panel of peptides in which you move the staple around to different positions and in some cases we’ve shown you have to tweak certain residues in order to maximise cell penetration, sometimes you need to redistribute residues on the peptide, sometimes you need to replace negatively charged residues with uncharged versions.’

Aileron’s first stapled peptide through Phase 1 clinical trials, ALRN-5281, boosts human growth hormone in patients with rare endocrine disorders. Vice president of clinical development, Hubert Chen explains: ‘Endocrinology and metabolic disorders are definitely going to be opened up by the stapled peptides approach, because of our ability to rationally design long-acting hormones’.

In this case, the drug’s target is on the cell surface. But the use of stapled peptides offers advantages over present therapies. Existing drugs provide artificial hormone replacements at dosages that lead to abnormally high hormone levels in patients, causing side effects such as swelling, joint pain and diabetes. The stapled peptide drug works by stimulating the body’s own growth hormone production. It is therefore better able to replicate normal physiological hormone levels and should reduce side effects

A growing number of studies show that stapled peptides can reach previously undruggable cancer targets. The peptides are able to turn on or off the processes that activate cell death or promote cell survival by disrupting how genes control protein synthesis.

In 2012, Aileron and Roche announced promising preclinical results on a stapled peptide cancer drug (ATSP-7041). The drug works by reactivating a cell death pathway known as p53. This pathway plays a critical role in protecting cells. ‘50% of all human cancers retain wild-type p53 and would be potential candidates for this mode of action – that’s an extraordinarily large number’, says Nash.

Stapled peptides drugs could also provide solutions to chemotherapy resistance, where previously successful cancer treatments suddenly stop working. Aileron is creating stapled peptides that tackle the B-Cell lymphoma 2 (Bcl-2) family of proteins, which are involved in cell death. Over-expression can lead to chemotherapy resistance, but stapled peptides are able to block their activity and activate cancer cell suicide pathways.

Another area studied is HIV infection. David Cowburn, at the Albert Einstein College of Medicine in New York and Asim Debnath at the New York Blood Center in the US, showed, in 2008, that a stapled peptide disrupts the viral outer shell or ‘capsid’, which will prevent replication. Cowburn concedes that the work is still at an early stage. ‘The affinity is still too low to use stapled peptide in their current published integration as a drug. The success of this scientifically is validating the mechanism fully,’ he says.

The development of stapled peptides has not been without drama. In December 2012, a paper from an Australian research team led by Peter Czabotar (ACS Chem Biol., 2013, 8, 2) received attention as it reported that several stapled peptides, originally designed in Walensky’s Harvard lab, did not lead to cancer cell death. Walensky responded that the stapled peptides tested were not designed as drugs but for use in NMR studies.

‘We weren’t surprised by the paper.’ Nash says. ‘The data were old news. The interpretation beyond that particular compound is not an accurate reflection of the state of the art.’

The controversy is a reminder of how new fields progress. In Verdine’s words: ‘There’s a field that’s being birthed here – that’s really the news.’ In an effort to speed up progress in the field, Verdine, together with Singapore-based collaborator David Lane, are planning to make all their published stapled peptide available to researchers. They want to ‘democratise stapled peptides,’ which are expensive to make.

But stapled peptides will not reach all drug targets and Verdine’s group is starting to work on second generation peptide based therapeutics that he calls ‘cell-penetrating mini-proteins’. They are learning how to stabilise mixed α and β structures.
Verdine points to other work in the general field of ‘constrained peptides’, where peptides are synthesised with cyclic portions to lock in biologically active structures.

In Switzerland, University of Zurich chemist, John Robinson has stabilised β-hairpins. Based on his work, Swiss start-up Polyphor has announced a new class of antibiotics that are effective against drug resistant bacteria.

Given the breadth of drug applications for stapled peptides, has progress been slow? Verdine, who is no longer associated with Aileron, comments: ‘Roche and Aileron are being, in my opinion, extraordinarily cautious. You can argue that this is good stewardship of the field, but for those of us who are really eager to see stapled peptides in patients, it’s taking a long time’.

Aileron acknowledges it has an embarrassment of riches. ‘There are too many good things for Aileron to work on all by ourselves,’ Nash comments. ‘We are in conversations and will hopefully develop other partnerships.’

Meanwhile, Verdine predicts that the full impact of stapled peptides is some time off. ‘I think its going to take a decade for there to be really significant numbers of approved molecules – we are really right at the startling line.’

Rachel Brazil is a freelance science writer based in London, UK

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