We continue our series on structural biology and structure-based drug discovery, and we will discuss screening this time. As I mentioned in the previous post, screening is central to any drug discovery strategy we may decide to choose. The screening aims to identify a compound or a series of compounds that can bind to the drug target and modify its activity. Before the start, we need to define two essential factors, the compound library we want to use and the screening method.

How to choose the compound library? We know that the chemical space is endless. However, the good news is that there is no need to have a library with billions of compounds. There are many ways by which we can limit the number of compounds for screening. One of the best-known is Lipinski’s rule of five (Lipinski et al., 2001) for orally available drug candidates. This is an empirical rule based on the analysis of known drug molecules. The rule states that an orally active drug should not have more than one violation of the following criteria:

  • Mw < 500 g/mol
  • Hydrogen bond acceptors < 10 (N and O)
  • hydrogen bond donors < 5 (OH and NH)
  • logP < 5 (logP=log ([Coctanol]/[Cwater])

logP is needed to ensure a certain level of solubility of the molecules. It describes the distribution of the compound in an octanol-water system, where [Coctanol] and [Cwater] are the concentrations of the compound in the octanol and the water phase, respectively. These conditions will impose tight limits on the potential number of compounds to be included in the library. We could further limit our choice by selecting compounds similar to known lead compounds (lead-like compounds) targeting a particular target class by defining shape complementarity to fit a specific binding site. Another essential factor is, of course, the chemistry of the compounds. They should be easy to modify chemically and should not contain atomic groups known to trigger a toxic response or unwanted reactions. Limiting the number of rotatable bonds (maximum seven) has also been an important parameter (Veber et al., 2002).

An alternative approach to using drug-like or lead-like compounds is fragment-based drug discovery (FBDD), which has gained considerable popularity due to its efficiency in identifying a pharmacophore. A pharmacophore describes the interactions that contribute to ligand binding and can be used in the design of new compounds during hit-to-lead optimization and the construction of a screening library. Screening is run using a relatively small library of molecular fragments with a molecular weight in the 100-300 Da range. The small size of the fragments will minimize the chances of unfavorable interactions like steric repulsion, increasing the probability of binding to the target protein and the hit rate. This will identify weakly potent and biologically active molecules. Complexes of these molecules with the target can be studied with X-ray crystallography or NMR spectroscopy. They will guide the hit-to-lead expansion and optimization of affinities and specificities into potent leads during hit-to-lead optimization.

A fragment library can be constructed to sample a much larger chemical space than lead-like or drug-like libraries. Another popular approach involves the creation of DNA-encoded libraries (DELs). DELs have the advantage of allowing an even larger chemical space coverage (for details, see, e.g., this summary and some reference publications).

For constructing a fragment library, the rule of three has been suggested (Congreve et al., 2003):

  • MW 100-300
  • LogP ≤ 3.0
  • H-Bond Acceptors ≤ 3
  • H-Bond Donors ≤ 3
  • Rotatable bonds ≤ 3
  • Polar Surface Area ≤ 60 Å2

While X-ray crystallography and NMR spectroscopy can be used for studying compound binding, they can also be used for library screening. Generally, in fragment screening, biophysical methods quickly became popular since they provide a rapid and efficient assessment of the binding of weak hits, which can be challenging to identify in biochemical assays. The most popular methods include the following:

  • X-ray crystallography
  • NMR spectroscopy
  • Surface plasmon resonance (SPR)
  • Thermal shift assay (differential scanning fluorimetry, DSF)
  • Weak affinity chromatography (WAC™)

SARomics Biostructures offers screening services using X-ray crystallography, NMR spectroscopy, and WAC™. WAC™ is a proprietary method jointly owned by SARomics Biostructures and our in-house partner Read Glead Discovery. The method uses a chromatography column into which a solution of fragments or lead-like compounds is injected. During elution, the fragments with a higher affinity for the protein will stay on the column longer than those with a low affinity. The fragments can be conveniently detected and identified by mass-spectrometry and thus establish a direct ranking of hits. WAC™ is an efficient, high-throughput, and lower-cost choice compared to other biophysical screening methods. In addition, we offer our proprietary library of MedChem-friendly low-molecular-weight fragments designed to be general-purpose (not target-directed), covering diverse chemical space. Of course, customers' libraries may also be used for screening. Several successful projects have already demonstrated the efficiency of the method.

We will continue this series of posts on structural biology and structure-based lead discovery and design. The following post will discuss how structural biology has contributed to drug discovery. Follow us on LinkedIn to ensure you do not miss our future posts!

Previous posts:

Why a structural biology services company?

Structure-Based Discovery Strategies