Move from hit to lead

A long-standing customer had a hit series with good activity but poor properties. Cresset Discovery Services worked closely with the customer to formulate a plan of action to optimize the compound properties while maintaining potency.

Cresset Discovery Services has worked closely with a customer on a target. We ran virtual screens, aligned literature chemotypes and proprietary chemotypes in order to arrive at a robust binding hypothesis and a field hypothesis. This enabled the customer to find a new chemotype that had good activity at the target. However, the hit series had poor properties. They needed to optimize the compounds, potentially sacrificing some potency while balancing this against improving the properties of the molecules.

The target in question required lipophilic molecules, so the set of compounds had reasonably high lipophilicity which can be a liability in drug development. The compounds also had high protein binding and, we suspected, high clearance.

 
Figure 1: Forge radial plot. The selected ‘highlighted’ set (Figure 3). Compounds with better properties give a larger area in the radial plot.

 
Figure 2: Forge graphical radial plot parameters. For each property a function is used to describe perfect, acceptable and unacceptable values. Perfect values are plotted at the edge of the radial plot and unacceptable plotted at the center. A single ‘Radial Plot Score’ is created to represent the fit of a compound to the chosen set as a function of parameters using the specified weighting scheme.

 
Figure 3: Forge property plots. Radial plot scores as a MPO method: Compounds with a radial plot score greater than the chosen cut-off (bottom left) were selected and hence are highlighted all the plots. These compounds have a good balance of activity and other physicochemical properties.

The best course of action for optimization was to search for compounds that were reasonably active but far more polar than the bulk of the molecules in the compound set. We designed suitable variants of these candidates and checked them using the alignment model in field space that we had developed earlier in the project. New combinations of functional groups on the core were selected to address the overall lipophilicity whilst maintaining the essential interaction features. This solution had the potential to address both the protein binding and clearance issues.

Over the course of the following weeks the customer worked through these ideas with a high degree of success. We collaborated closely with the customer throughout the project. They shared not only their activity data, but also their property data. We assessed both the activity and property landscape, and refined the suggested sets with the aid of multi-parameter optimization, which enabled us to suggest which compounds they should make and progress to help to get them past the hurdles they were experiencing. The customer has now arrived at a set of compounds with far better properties without a significant loss of potency.

The customer has retained our services to help them to optimize the potencies of the back-up series, which may help them to choose which chemotypes to progress next.

Contact us to see how we can help you with similar projects.

Conduct ligand-protein docking

A long-standing customer of Cresset Discovery Services asked us to identify new compounds that could be active at their protein target. We conducted ligand-protein docking to narrow down their 50k compound library to the best 1.5k compounds. The cost of the consulting project plus the chemistry for 1.5k compounds was about 20% of what it would have cost to buy and screen the entire 50k library.

Ligand-protein docking can be an excellent way to build up knowledge about the binding pocket. It can also form the basis for a virtual screen to identify new active compounds.

Cresset Discovery Services had been working with this customer on a particular ligand for some time, but there was very little information available about the protein target. There were homologues in the literature, but they were distantly related and nothing very similar had been crystallized.

Detailed preparatory work to model the protein active site

It was necessary to do a lot of modeling work to build up the relationship between the human target and the distantly related proteins available from the literature. We built sequence alignments and compared them, enabling us to build up 3D models of the target and its interaction with the ligand.

Some mutagenesis data was available on the known ligands, so we were able to use this to refine the 3D models and check that the correct residues were in the right places on the active site. This enabled us to define the active site for the ligands. We went on to calculate the energies for the protein-ligand interactions to make sure we had identified poses that made sense.

This was a complex system that required a great deal of protein preparation. This preparatory work was essential for successful docking and required expert knowledge, experience and skill.

Docking and virtual screening using different scenarios

At the end of this process we had a good model of the protein-ligand system. The next step was to remove the ligand and carry out docking.

Docking was first tested on the molecules that were known to bind to the target. This resulted in excellent retrieval rates, showing that the model would also be able to retrieve new compounds.

There were a number of different binding sites on the protein so we decided to carry out the virtual screening using different scenarios for the protein. We:

  • Kept the ligand intact in the binding site
  • Removed the ligand completely
  • Looked at partly bound situations and un-bound situations for each of the binding sites.

The customer provided us with a set of 50k ligands and we docked each of these against the binding pockets. A docking scoring system was used to rank the top 2k compounds from each of the screens.

Analyzing the results and compiling a purchasing list

The top 2k compounds from the four screens were analysed in detail. We visualized every one of the top 2k compounds and looked at each of the docking poses. The docking gave us good geometries for the ligands and we used Cresset software to check that the electrostatics made sense. Any compounds that were unlikely to bind well were rejected.

A final, ranked list was provided to the customer with a very high degree of confidence that it included compounds that were active at the protein target. They were able to procure about 75% of the compounds from the hit list, giving them a final set of 1.5k compounds to test.

An incredible saving in time and money

Carrying out virtual screening to focus the library in this way represented an incredible saving in time and money for our customer. The alternative approach would have been to buy and test the whole 50k compound set. Not only would the customer have needed to purchase all of the compounds, but also shipped them, stored them, plated them, screened them, and then they would still have to analyse the results.

The estimated cost of doing this for all 50k compounds would have been about five times the cost of the combined tasks of the Cresset Discovery Services project plus buying and testing 1.5k compounds.

Cost of

{buying and testing 50k compounds}

=  5 X

Cost of

{Cresset Discovery Services project + buying and testing 1.5k hit list}

Contact us to find out how we can add value to your project.

 

 

 

 

 

Dr Martin Slater, Director of Consulting Services

Cresset Discovery Services case studies

Cresset Discovery Services manages and delivers outsourced computational chemistry programs. We work alongside chemists to solve problems, provide fresh ideas, remove roadblocks and add direction and insight. We work on:

  • Lead generation and optimization
  • Creating, broadening or protecting IP
  • Supporting grant funding applications
  • Accelerating the commercialisation of assets.

What we actually do depends on the customer project, which is usually confidential. Our case studies give an idea of the breadth and flexibility of our work and include some anonymized examples of customer projects.

Homology modeling and ligand electrostatics plays key role in elucidating binding mode and molecular interaction of new class of antifungal drugs

Last month F2G published a paper in PNAS [1] describing F901318, the leading representative of a novel class of antifungal drug. Dr Martin Slater, Director of Cresset Discovery Services, is a co-author on the paper. He describes how modeling work carried out by Cresset Discovery Services was critical to predicting the binding mode of the inhibitor and important interacting amino acid residues. F901318 is currently in clinical development for the treatment of invasive aspergillosis.

There is an important medical need for new antifungal agents with novel mechanisms of action to treat the increasing number of patients with life-threatening systemic fungal disease and to overcome the growing problem of resistance to current therapies.

F2G are a UK-based antifungal drug discovery and development company who have identified F901318 as a leading representative of the orotomides, a novel class of antifungal drug. Their identification of dihydroorotate dehydrogenase (DHODH) as the mechanism by which F901318 inhibits and kills Aspergillus fumigatus has been a major breakthrough differentiating F901318 from other systemic antifungal agents.

From hit to lead with medicinal chemistry

F2G had a large amount of proprietary cellular activity data developed over time against their antifungal screening platform. After an initial hit finding campaign significant progress had been made using classical medicinal chemistry approaches.

F2G were keen to inform and assist the development process by gaining a molecular level understanding of the target protein ligand system. They approached Cresset Discovery Services for help in elucidating the molecular interaction of the target protein-ligand system.

A detailed molecular understanding with modeling

Cresset’s unique approach of defining the electrostatics around the active chemotype made it possible to identify the precise nature of the various sites on the active molecules. In conjunction with sequence analysis across the wider DHODH family, Cresset scientists were able to match these subtle ligand features to the patterns of residues that were likely to be key.

Subsequent homology and ligand protein interaction modeling of Aspergillus fumigatus DHODH using the XED force field identified a predicted binding mode of the inhibitor and important interacting amino acid residues.

We combined a detailed ligand centric approach using Forge with protein modeling using a prototype of the new Cresset protein tool to arrive at a binding hypothesis consistent with the selectivity profile. The modeling process is fully reported in the paper [1].

Testing in silico hypotheses in vitro

Having made a binding hypothesis, a number of lab experiments were initiated by F2G to check the predictions e.g., using site directed mutagenesis.

Most satisfyingly, the lab results supported our predictions.

F901318 is currently in late Phase 1 clinical trials, offering hope that the antifungal armamentarium can be expanded to include a class of agent with a mechanism of action distinct from currently marketed antifungals.

Cresset’s consulting work with F2G provided valuable insight into the predicted interaction pattern of the main chemical series with the Aspergillus DHODH target protein. As with many research projects, any level of understanding achieved is often a prelude to even deeper questions, and there are many remaining to be answered for this unique system. Cresset continues to work closely with F2G, providing software and services to support them in their ongoing projects.

References

1 http://www.pnas.org/content/113/45/12809.abstract

 

 

 

 

 

 

 

Dr Martin Slater

Director, Cresset Discovery Services

Build and cluster diverse 3D libraries

Cresset Discovery Services (CDS) worked with BioBlocks to analyze their fragment library to maximize coverage of 3D chemical space. As part of the project, we developed an innovative clustering method that made it possible to assess the 3D similarity across their virtual database of over 1.5 million fragments.

The goal of the project was to help BioBlocks build the maximum 3D diversity into a fragment library of manageable size from a starting pool of over a million compounds. Existing techniques would have required an infeasible amount of computing power, so CDS developed an entirely novel rapid clustering method especially for the project. The solution was still extremely computationally challenging, but we were able to use our expertise in distributing calculations to the cloud to deliver the results that BioBlocks needed on time and within budget.

“Working with Cresset has been a positive experience from start to finish,” said Warren Wade, VP of Chemistry at BioBlocks. “Because our fragments are designed to be new chemical matter, they challenged the limits of existing structural descriptions. Cresset worked closely with us to overcome these limits and produce a high value compound set”.

The final result was a 3D fragment library that contains a significant number of compounds with novel core structures that are now viable candidates for fragment screening. BioBlocks envisions this Comprehensive Fragment Library to be a drug discovery tool available only to collaborators who will be able to leverage this new chemical space for their lead discovery programs. Hits from the library are entry points to BioBlocks’ collaborative medicinal chemistry processes, developed to increase the probability of generating commercially viable leads.


3D Similarity-based clustering workflow
3D similarity-based clustering workflow

Read more about this project: Large scale compound clustering in 3D.

Contact Cresset Discovery Services to find out more about how we can help you design large scale libraries for your project.

Develop bespoke software

Cresset software focuses on novel methods to discover, design, perfect or view compounds and their data in easy to use applications. Our applications are firmly founded in the experience of our customers and the most common problems that they face. However, our development expertise is not limited by our applications. We have an excellent track record of delivering novel scientific plugins, command line applications and workflows that go beyond our commercial offerings.

Custom integration

Many customers have extensive in-house computational chemistry tools that their chemists appreciate and derive value from. They like to access the unique benefits of Cresset software from within their existing solutions. In this situation Cresset Discovery Services (CDS) can develop software that is seamlessly integrated with the customer’s existing framework. From viewing our excellent electrostatic interaction potentials to detecting 3D activity cliffs we have the expertise to plug Cresset directly into their world.

Unique science

Customers regularly approach us with specific workflows, or novel approaches for which they require a truly bespoke solution. We can either develop bespoke software with an exclusive license, or co-develop novel methods for the customer to their requirements, before developing it for a wider commercial market. In either case we use our extensive experience of creating software for chemistry workflows to deliver usable, stable and functional solutions that solve the customer needs.

“Bespoke software gives customers access to Cresset software without having to change their software environment.”
Dr Martin Slater, Director, Cresset Discovery Services

Case study

Cresset were recently asked to develop a tool that could look at a customer’s SAR data and prioritize new molecules for synthesis based on the information that they would add to the project, as well as potential for activity. The outcome of this work was a ground-breaking 3D-QSAR application. As part of the customer agreement, Cresset went on to commercialize the application as Activity Atlas, which is now a component of Forge.

 

If you have a situation that requires a novel solution, get in touch for a free, confidential discussion to find out how Cresset Discovery Services can work with you to develop a bespoke application.

What’s in the CDS virtual screening toolbox?

Cresset is very well known for providing fast and accurate ligand-based virtual screening through Blaze. We have now added the Lead Finder docking engine to our virtual screening toolbox, giving Cresset Discovery Services (CDS) the most comprehensive virtual screening capabilities available anywhere in the industry.

Based on an informal survey of our contacts and customers, I estimate that something like 50% of all current pharma SME projects are ‘structure enabled’. Lead discovery and lead optimization are driven through the use of in-house structures, public structures (typically from the PDB) and homology models. These structures inform lead optimization programs by explaining observed SAR and providing feedback and a detailed context for the design of further analogues.

CDS routinely uses the Cresset software Blaze for ligand-based virtual screening. Although we had access to structure-based methods, we are pleased to have brought Lead Finder in-house, giving us full capability in conducting ligand-protein docking.

Ligand-based virtual screening with Blaze

Virtual screening with Blaze remains one of the most consistently requested projects for CDS. What makes Blaze extremely useful for our customers is:

  • Virtual screening is probably the only way to really sample adequate chemical diversity
  • Virtual screens are far more cost effective than wet HTS
  • Excellent enrichments can be achieved
  • The chemotype diversity in the output is second to none.

Blaze also relies on two very simple premises:

  1. A bioactive conformation encodes, in its shape and electrostatic field, both the properties, recognition features and solvation pattern optimised for interaction with its protein target site.
  2. A molecule conformation with increasing ‘shape and field’ similarity to that bioactive conformation has an increasing probability of also being active.

So, the key determinants of real activity obtained from hit lists (other than was this truly the ‘bioactive conformation’?) is often just how relevant and what distribution that hit conformation has in the population. This is fundamentally why our ligand-centric screening invariably works extremely well. Given that a molecule can adopt a similar shape, and project the same electrostatic patterns, from a completely different chemical architecture, leads to a very diverse output.

Structure-based virtual screening with Lead Finder

The Lead Finder software has been developed to provide cutting-edge docking for an array of typical tasks, from high-throughput virtual screening to best-in-class prediction of bioactive conformations to accurate prediction of binding energies. In combination with the companion Build Model protein preparation tool, Lead Finder has been shown to match or outperform the historically leading docking solutions.

When preparing ligands for virtual screening in Blaze, CDS scientists use modeling to help define the best ‘hand-crafted’ estimate of a bioactive conformation, based on the widest data for any given system. We apply the same care to exploring and preparing protein targets prior to structure-based virtual screens. We take advantage of three main approaches. Firstly, Lead Finder includes the excellent Build Model protein preparation tool. Secondly, we are privileged to be able to model proteins and ligands using the same proprietary XED force field used to give the accurate electrostatics that all Cresset software is based on. Finally, at CDS we have access to the latest Cresset software that is still under development. This gives us capability to provide protein electrostatic field maps and water analysis, providing a very reliable starting position for structure-based virtual screening.

vs_2bsm3

Lead Finder uses a stochastic ligand sampling workflow, with conformations generated on-the-fly, and a genetic algorithm for processing these into pools of the best docking poses. Multiple interaction grids are generated from the protein target and combined to define a scoring system for poses. More importantly, the scoring method has been shown to outperform some of the more conventional docking engines currently available commercially.

Structure-based or ligand-based?

What are the advantages of having structure-based and ligand-based virtual screening?  And how do we choose which is the best approach for a project?

Ligand-based virtual screening is less computationally intensive, making it a preferred option when there is a known ligand available. An average protein of 400 amino acids has over 20,000 heavy atoms and 9,600 bonds and in excess of 50 charges, making it a more challenging system to model.

However, even when there is a known ligand there are some situations when a ligand-based virtual screening is not viable, such as when the known ligand does not exploit all the interactions available in an active site or when a protein has an unattractive orthosteric site and attractive allosteric sites with no known ligands. In these cases, we prefer to use a structure-based method.

In the case of protein-protein interaction sites and protein-DNA/RNA sites, Blaze can take DNA and protein fragments as a template in place of a ligand. However, it is useful to have a structure-based approach available for comparison.

In fact, we often find it useful to combine different virtual screening techniques. In lead discovery, one of the key requirements for virtual screening is to maximise the diversity of hits returned.  All virtual screening techniques, be they ligand-based or structure-based, are probabilistic techniques in that they may be used to increase the likelihood of getting hits from a wet screen. No technique guarantees to give absolute binding energies (at least not in the context of virtual screening on any realistic size of screening library), but they do give good rank ordering of compounds and can, therefore, be used as a means of selection and prioritisation.

Ligand-based techniques, whether 2D or 3D, are algorithmically distinct from structure-based techniques such as docking and, therefore, give different rankings to compounds. Different approaches return different hits and the results can be combined into an enriched final list.

Combining the results of structure-based and ligand-based techniques provides further diversity, leading to better hit rates and more interesting hits.

A one-stop shop for virtual screening

Through combining the strengths of Blaze in the ligand-based world with Lead Finder for docking, CDS now has the most comprehensive virtual screening capabilities available anywhere in the industry. Both Blaze and Lead Finder are available to purchase as software or as a service through CDS. CDS is truly now a one stop shop for virtual screening and indeed very much more.

Download a free evaluation of Lead Finder or access the Blaze demo server.

A new breed of CRO

Cresset Discovery Services (CDS) is a new breed of CRO. This is evident in two core principles that guide our work.

The first is to use excellent in silico methods to aid small molecule discovery and design.

We believe that computational chemistry tools provide intelligent short cuts, enabling us to select ‘20% of compounds that count’ to progress to the next stage of design or optimization.

Computational techniques also give an extra level of insight to chemistry and biology. In the absence of a crystal structure, computational methods are the best way of deducing information about the protein target and the likely active conformation of the ligand.

Unlike traditional serial and parallel techniques, computational design allows for iterations, meaning that conclusions can influence thinking and the next stage of experimental design.

A new breed of CROThe second principle that informs our working style is our partnership approach. Our expertise is in silico work and we create synergy by working with others who are experts in their own field. While we can recommend partners, you can also ask us to act as the project manager – sub-contracting work such as procurement, X-ray crystallography, modeling and synthesis to our partners. This makes the process seamless for you as you then just have one point of contact and one invoice to process. The following examples demonstrate how this works in practice.

Delivering plated compounds

The results of a virtual screening experiment are a list of compounds to be purchased for biochemical screening. Rather than receiving a list of potential compounds to purchase, you can ask us to take the next step and carry out the procurement for you.

We have preferred suppliers for procurement and the delivery of plates. This gives you one point of contact, saves you the logistical headache of working with multiple suppliers and takes care of any customs considerations for you.

More than that, it means that we can use our insight into the results to make the best possible substitutions if a compound is unavailable or out of your budget. Further information: Managing the procurement process to deliver plated compounds.

Chemistry and crystallography

A current project involves working out how the 3D bioactive conformation of active molecules interacts with the target protein. Essentially, we are doing some protein detective work. CDS have carried out some X-ray crystallography to check the molecules the customer had. We will then go on to design new molecules, get them made, do the crystallography and get the chemistry done with our preferred suppliers and partners.

Our chain of partners means that we can manage your discovery process from target validation and identification, finding a molecule, synthesising it, getting it tested and doing first stage trials.

Contact us today to work with Cresset Discovery Services: A new breed of CRO.

Engaging with Cresset Discovery Services

Cresset Discovery Services (CDS) offers bespoke in silico services for small molecule discovery. We do a lot of work in drug discovery and optimization for the pharmaceutical industry but we also work extensively in agrochemicals, flavors, fragrances – in fact, in any industry that involves work with small organic molecules.

This post explains the process that we go through when customers work with Cresset Discovery Services, from the first contact to the final deliverables.

Enquire

At the enquiry stage we talk with customers about their requirements in general terms to get an idea of whether we will be able to help them. The answer is usually yes, but we will certainly let you know if we think that our approach would not be the best match for your project.

These initial discussions will involve members of both our sales team and the scientific team. Everything at the enquiry stage is free, but the discussions will not be at a great depth since confidential details cannot yet be shared.

Establish

Once both sides have agreed to proceed, we exchange confidentiality agreements and can then get down to the details. The customer will share their confidential data and CDS will prepare a detailed proposal of the work they will carry out.

This stage will involve a detailed meeting to gather the data and another to present the proposal. The proposal will include full details of pricing and milestones. If the work is a collaboration, then all partners will be involved at this stage.

Execute

Close collaboration is key to any successful project. Depending on the size and complexity of the project, there may be several long meetings at the start of the project. These could involve many members of the customer team. The goal is to focus on the project and to scope out exactly what needs to be achieved.

Work then moves to the details – for example, what to do, with which molecules and which conformations. This could involve conversations several times a week until everything is in place to run the study.

Frequent reviews take place throughput the project between the customer and CDS. Each customer has a personal point of contact who remains consistent throughout the project.

At each stage of the project there will be several conversations to make sure that the customer is getting exactly what they wanted. These will be tied in to agreed milestone reviews and deliverables.

Project deliverables are likely to be available through the project, not only at the end. No matter when they are delivered, the approach remains the same: we make sure that the customer gets the maximum value out of the results.

For example, typical results for a large screening project with multiple compounds may be between 10,000 and 20,000 hits. But CDS will make sure that the customer gets more than a list from the project. We will always ensure that the customer fully understands and can interpret the results in the context of the project in order to get the best out of them.

Evaluate

No project is complete without a project review of what went well and what could go better. As part of this process we agree the next steps, which could range from a follow-on project, to advice on the next research steps.

Many of our customers remain customers for the long term. In fact, when we do lose a services customer it’s usually because they have decided to buy our software and hire a computational chemist to work full time. This case study describes how we helped one customer to hire and train computational chemists. Even then, customers still come back to us for projects if they need the extra resource.

 

Contact us today to start the process of working with CDS.

Enquire_Establish_Execute_Evaluate

Identifying possible protein targets

An academic group had identified a molecule that was active in a phenotypic assay. Due to its similarity to other compounds, they initially thought it was acting through a known pathway. However, further work revealed it was acting through a different pathway. The challenge was to identify the protein target. They chose a computational method and outsourced the project to Cresset Discovery Services (CDS).

A computational approach to identifying protein targets

There are a number of ways to go about identifying possible protein targets. The biological route is the standard method of trying to de-convolute the target from the cellular context. However, it involves a lot of time and work. Taking a computational approach can be a very efficient shortcut that narrows the search down to a few likely candidates.

The academic group choose to take the computational route and they decided to outsource this sophisticated project to CDS. Cresset was the ideal choice for a number of reasons.

The cost and time advantages of outsourcing

Firstly, it was far quicker and cost effective to outsource the project rather than trying to do it in-house. To do the work in-house would have needed to recruit an expert and to purchase the software and hardware. There would have been steep learning curves for using the software and methods. Added to this, they would potentially have been left with these overheads at the end of the project.

By contrast, for an outsourced project CDS supplies both the expertise and the infrastructure. This makes it far quicker to get up and running. It is also far more cost effective since the resources are only paid for the duration of the project.

Choosing partners who have the right expertise

Added to these advantages, CDS has specific expertise in this field. Our software calculates the interaction potentials of molecules by analysing their shape, electrostatic fields and hydrophobicity relative to atomic probes. These values are used to create a unique representation of

“This outsourced project was a very cost effective way of cutting down the list of potential targets”
 

molecules, making it possible to build up a ‘protein’s eye view’ of a compound. This gives a quantitative basis for comparing compounds based on how they are likely to interact with a protein.

We had already encoded the PDB ligands into Cresset field space, which translated to a head start in identifying the possible protein targets.

Carrying out the work

CDS took the active molecule as a starting point and found a number of conformations for it. We used the field patterns to search the PDB for similar molecules in order to find potential pharmacological matches. These matches revealed which proteins were worth investigating further.

From this work, CDS provided a list of potential targets to the customer. The academics worked through this short list in order to identify the actual target. This outsourced project was a very cost effective way of cutting down the list of potential targets and was certainly far more efficient than taking a purely biological route.

 

Contact us today to find out how Cresset Discovery Services can use their expertise to help with new approaches for your project.