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Drug developers have been in a quandary for some years now. The Human Genome Project  offered novel insights into disease, including numerous tantalizing targets for therapeutic intervention. Unfortunately, many of these are protein-protein interactions (PPIs) not readily addressed by conventional “Rule of 5” small-molecule drugs. The protein surfaces are too large, flat and poorly functionalized to be bound by small-molecules. Instead, protein therapeutics—such as antibodies and soluble receptors—have revolutionized the treatment of diseases such as rheumatoid arthritis, osteoporosis, psoriasis, and cancer.

Despite their therapeutic potential, proteins cannot enter cells and are not orally bioavailable. Large numbers of intracellular PPI targets remain stubbornly resistant to conventional drug discovery. Furthermore, where biologics are available, they are administered by injection, a painful and inconvenient process for the patient, especially for chronic conditions that require years of therapy.

In order to fill this unmet need, a number of companies—including Ensemble Therapeutics, Tranzyme, Aileron, and Bicycle Therapeutics—have started to investigate macrocycles. Macrocycles are small molecules that can act like biologics but are orally bioavailable and can also access intracellular protein-protein interaction targets. In fact, partnership activity for macrocycle-focused companies has surged in the last five years due to increased demand and significant potential value for oral biologics.1

Ensemble Therapeutics is creating macrocycles that have sufficient size and make enough interactions to disrupt even the most refractory PPI, but are lithe enough to squeeze through cell membranes. As such they merit the description “small-molecule oral biologics” and present a radically new drug modality.

The power of this approach has been demonstrated with the discovery of novel interleukin-17 (IL17) antagonists. IL17 is a cytokine central to a number of inflammatory diseases, such as psoriasis, psoriatic arthritis, and ankylosing spondylitis. Several biologics are currently in clinical development targeting IL17 and have shown stunning activity against psoriasis and other inflammatory diseases. However until now, there have been no small-molecules that bind IL17 – indeed most researchers would argue that IL17 is a target that would not yield small-molecule ligands. Macrocycles discovered at Ensemble bind to IL17 down to picomolar level, and have oral bioavailability and efficacy in inflammatory disease models.

The chemical technology platform (DNA-programmed chemistry) on which Ensemble Therapeutics was founded, employs DNA in a biomimetic fashion to drive chemical reactions. Macrocycles made by this method have a covalently bound oligonucleotide that provides a decoding tag for compounds active in affinity selection screens. The technology is routinely used to make millions of novel macrocycles, and screening these libraries has given hits against disease-relevant targets. In 2011, Ensemble discovered cytokine IL17 antagonist molecules in the micromolar affinity range. Methodical medicinal chemistry resulted in a thousand-fold affinity improvement and introduction of drug-like properties such as solubility, metabolic stability, and permeability. With proven efficacy in acute and chronic animal inflammatory models, a pre-clinical development candidate is anticipated in 2013.

Using macrocycles as small-molecule biologics          
Natural product macrocycles with pharmacological activity, such as cyclosporine and rapamycin, are a testament to the importance of this class for binding to protein surfaces. Many are considerably smaller than biologics with molecular weights in the 500-1500 Da range. However, they are only available in small amounts from natural sources and their structural complexity makes then difficult to synthesize or modify.

By creating millions of synthetic macrocycles, companies can generate novel hit compounds against diverse PPI targets. Rapid follow-up by milligram-scale synthesis permits confirmation of target affinity and functional activity, plus secondary cell-assay screening and pharmacokinetic profiling. Unlike macrocyclic natural products, these compounds are readily synthesized allowing potency optimization and fine-tuning drug-like properties, providing leads for previously inaccessible targets.

For IL17, two macrocycle classes have been optimized at Ensemble and are nearing nomination stage following numerous improvements over the original library hits. Binding affinity has improved 1,000-fold with simultaneous achievement of gut absorption and metabolic stability providing outstanding pharmacological efficacy in multiple disease models.

Many macrocycles possess binding kinetics similar to antibodies. The IL17 antagonists are potent binders with picomolar Kd values, and have fast on-rates and exceptionally slow off-rates. Macrocycles make many individual binding interactions with the protein—just like a biologic—resulting in a dissociation half-life that can stretch for hours, resulting in extended PPI inhibition with beneficial pharmacodynamics.

Synthetic macrocycles epitomize small-molecule biologics, presenting attractive aspects of both conventional small-molecules and biologics. They have proven affinity for challenging PPIs and yet retain drug-like small-molecule properties including cell penetration and oral bioavailability. In addition, the antibody-like binding kinetics confer exceptional potency and duration of action. Companies are refining methods to generate millions of macrocycles for screening and in Ensemble’s case have applied this approach to the discovery of small-molecule IL17 antagonists.

1. Cain C. BioCentury 20(38), A7–A13; Sept. 17, 2012.