Edward H. Kerns

Chapter 7 – Solubility

Publisher Summary nSolubility is one of the most important properties in drug discovery. Insoluble compounds can plague discovery. Solubility problems can intensify during discovery because the molecular characteristics needed for strong binding to the target protein can be deleterious to solubility. Solubility is determined by many factors, namely, composition and physical conditions of solvent, types of solvents, amount (%) of co-solvents, solution components, pH, and temperature. In drug discovery, various solubility experiments are performed to estimate the effect of solubility in different systems to better mimic the actual in vitro and in vivo conditions. Solubility is affected by physicochemical properties, which can be estimated using in vitro assays or software calculations. Medicinal chemists have the ability to change solubility by modifying the structure, which affects these physicochemical properties. Salt forms typically are selected to modify physicochemical properties such as dissolution rate, crystallinity, hygroscopicity, and mechanical properties (hardness, elasticity, etc.), leading to increased bioavailability, stability, and manufacturability. Examples of commercial drugs and their salt forms are summarized.

Plasma Protein Binding

Plasma proteins can adsorb a significant percentage of drug molecules. Binding to plasma protein can affect the pharmacokinetics (PK) of the drug substance in tissues and blood as well as the dosing regimen for the drug product. Drug compound molecules that are dissolved in the blood are in equilibrium with plasma proteins. The interaction of drug molecules with plasma proteins is electrostatic and hydrophobic. Plasma protein binding (PPB) is reversible. It is important to determine PPB across different species to establish safety margins for human exposure and doses for clinical trials. PPB impacts the PK of a drug and exposure to the therapeutic target. This chapter discusses various PPB effects so that drug discovery project teams can consider all of the possibilities. Quantitative structure–activity relationship studies have evaluated the physicochemical properties that correlate with PPB. PPB can be useful, retrospectively, as part of an ensemble of in vitro diagnostic tests to understand the impact of PPB on PK or pharmacological effects.

Plasma Stability Methods

Testing for stability in plasma or blood is advisable under certain situations (e.g., labile substructure, high in vivo clearance, prodrug research, unstable bioconjugate, and peptide). Compounds are incubated with plasma in vitro to predict in vivo stability. Analytical formats from single sample to high-throughput microplate procedures can be used. Liquid chromatography/mass spectrometry (LC/MS) is used for quantitation owing to sensitivity and selectivity with the complex plasma samples. Identification of degradation products using LC/MS techniques indicates the hydrolytically labile sites, so that structural modification can be tried to reduce the instability.

Blood-Brain Barrier Methods

Many tools are used to provide data for blood-brain barrier (BBB) permeability, brain exposure, and unbound blood to brain distribution. The data are used to guide brain exposure optimization, develop pharmacokinetic-pharmacodynamic relationships, and plan clinical studies. Peripheral drug discovery teams use the data to guide BBB permeation reduction to minimize central nervous system side effects. Passive transcellular diffusion BBB permeability ( P app ) is assessed computationally using logxa0 P and topological polar surface area and in vitro using parallel artificial membrane permeability assay-blood-brain barrier, Δlogxa0 P , immobilized artificial membrane, high-performance liquid chromatography, and Madin-Darby canine kidney cell monolayer permeability. Efflux ratio is assessed using in vitro MDR1 (multidrug resistant protein 1)-MDCKII (and other cell lines transfected with specific human BBB efflux transporter genes) and Caco-2, and in vivo using mouse strains knocked out for specific BBB efflux transporters. in vivo neuroPK studies provide an array of BBB permeability, transporter effects, brain exposure data ( C bu , AUC bu ), and unbound brain to blood partition coefficient ( K puu ). Brain binding studies, using equilibrium dialysis of brain homogenate or brain slice method, provide f ub , which is used to convert in vivo total brain concentration to C bu . Microdialysis and cerebrospinal fluid (CSF) sampling in vivo directly measure C bu and CSF concentration.

Chapter 6 – pKa

Publisher Summary nMedicinal chemists can modify the acidic or basic substructures on the scaffold in order to obtain the desired pKa, which affects solubility and permeability. The ionizability of a compound is indicated by pKa. Ionized molecules are more soluble in aqueous media than neutral molecules because they are more polar. Solubility is determined by both the intrinsic solubility of the neutral molecule and the solubility of the ionized species, which is much greater. The effects of ionization suggest a relationship frequently encountered by medicinal chemists: highly permeable compounds often have low solubility and vice versa. Thus, there is a tradeoff between solubility and permeability because of the opposite effects of ionization on these properties. Examples of the pKa values of a number of substructures that commonly appear in drug molecules are listed. It also presents examples of the effect of pKa and molecular size on the activity of a structural series, the effect of pKa on activity, and the effect of pKa on water solubility. By modifying the substructures of a molecule to introduce groups with differing pKa values, medicinal chemists can modify the solubility and permeability of the compound.

Chapter 5 – Lipophilicity

Publisher Summary nLipophilicity is a property that has a major effect on absorption, distribution, metabolism, excretion, and toxicity (ADME/Tox) properties as well as pharmacological activity. Lipophilicity has been studied and applied as an important drug property for decades. It can be quickly measured or calculated and is correlated to many other properties, such as solubility, permeability, metabolism, toxicity, protein binding, and distribution. The traditional approach for assessing lipophilicity is to partition the compound between immiscible nonpolar and polar liquid phases. Lipophilicity is an underlying structural property that affects higher-level physicochemical and biochemical properties. It changes with the conditions of the phases, including the following: partitioning solvents/phases, pH, ionic strength, buffer, and co-solutes or co-solvents. It can be correlated to various models of drug properties affecting ADME/Tox. They include permeability, absorption, distribution, plasma protein binding, metabolism, elimination, and toxicity. It often is an effective guide for modifying the structure of a lead series to improve a property. The effects of lipophilicity on specific properties and structure modification strategies are discussed.

Metabolic Stability Methods

It is difficult to predict metabolic rate using software, but predicting the labile sites is more successful. Various combinations of subcellular fractions with cofactors and hepatocytes are used to study metabolism in vitro. The different systems cover different metabolic enzymes. Structure elucidation of major metabolites guides structure modification to increase stability.

Strategies for Integrating Drug-like Properties into Drug Discovery

How and when property data are used can have a great impact on a project. Successful property strategies include the following: start assessing properties early, profile properties of new compounds rapidly, develop structure-property relationships, optimize activity and properties in parallel, use single-property assays to guide specific modifications, improve bioassays and their interpretation, customize assays for specific research questions, diagnose the root cause of pharmacokinetics liabilities, and use human materials to predict human performance.

Plasma Protein Binding Methods

The importance of free (unbound) drug for distribution into disease tissues and clearance organs has prompted many drug companies to implement plasma protein binding (PPB) measurement methods. Literature and commercial in silico predictors of plasma protein binding are available. This chapter discusses various in silico models for PPB. PPB is found to increase with increasing lipophilicity, increasing acidity, and increasing number of acid moieties. PPB decreases with increasing basicity and number of basic substructures. Several commercial software products for predicting PPB are available. In vitro PPB methods are also reviewed. Concomitant studies of PPB using more than one method have indicated the comparability of results from several other methods. The methods discussed are equilibrium dialysis, ultrafiltration, ultracentrifugation method, and immobilized protein high-performance liquid chromatography column method. Many other methods for analysis of drug–protein complexes have been reported. Some methods measure specific properties of the drug–protein complex: fluorescence spectroscopy, circular dichroism/optical rotatory dispersion (CD/ORD), NMR, electron spin resonance (ESR), microcalorimetry, and surface plasmon resonance.

Lead-like Compounds

Lead-like compounds have lower initial values for structural properties, allowing increases without becoming non-drug-like. The “hits” that serve as starting places for leads come from high-throughput screening, virtual screening, natural ligands, natural products, and the scientific literature. In the “hit-to-lead” phase, it is important to include properties in the workflow and goals for lead selection. In this evaluation process, some effective concepts have been emerging: lead-likeness, template conservation, triage, and fragment-based screening. The use of one or more of these concepts can increase the chances of success in discovering a strong lead-like structural foundation. Structure modifications during optimization are often added onto the lead template, thus retaining much of the original core structure of the lead. An emerging strategy in exploring for novel leads is termed as fragment-based screening. The well-studied lead-like criteria reinforce the principle that success in discovering drug-like clinical candidates is higher when starting with compounds that have structural properties with lower values, which allow the opportunity for structural modifications to enhance activity and selectivity without condemning the series to unacceptable pharmacokinetic performance.