Structure-Activity Relationship Studies in Drug Development by NMR Spectroscopy

The aim of pharmaceutical research in the present scenario is to modulate the targets with the intention of evoking therapeutic benefits. The advent of DNA recombinant technology has facilitated the preparation of almost any type of target in the purest form and in required amounts. In parallel, combinatorial chemistry and high throughput organic synthetic approaches have dramatically expanded the number of compounds that can be evaluated for biological activity. Both these advances have paved the way to develop high throughput screening systems for evaluation of millions of compounds against these targets in a rapid and efficient manner. However, overall return on investment in these technologies has been meager and disappointing. Thus an alternate approach which has the ability to potently and specifically modulate protein targets with small organic molecules is a need of the hour. All these facts have contributed to the emergence of multidimensional NMR as a gate way to drug design and development.

Fragment based drug design is the concept, where ligand binding site is dissected into different regions and the binding of diverse compounds in individual sub sites are analyzed separately. Fragments with fairly good affinity at all sub sites are collected and are interlinked in various possible manners, intuitively to generate a new set of compounds. The affinity of these compounds will be the sum of the affinity of individual fragments with additional gains due to entropy. In this approach, ligand-target interaction is measured and characterized with at most accuracy and precision as it caters valuable information in the areas like quantitative determination of binding affinities of potential ligands, ligand binding site determination on protein, conformation information of ligands and proteins and local flexibilities in the structure for complex and free components. In addition NMR helps to determine ligand specificity and selectivity towards different targets and also provide valuable hints to predict possible side effects and toxicological outcomes at various levels. Another advantage of these techniques includes attaining capability to eliminate the false positives. They are also useful in further optimization of the potential leads.

Drug receptor interaction is quantified based on various techniques through NOE, relaxation and diffusion editing, isotope editing/ filtering, shapes strategy, HSQC strategy, etc. Fragment linking and elaboration strategies are employed for linking the fragments together and design the new molecules. Enormous number of compounds with high affinity towards several targets from FKBP to Bcl-2 has been reported in the literature.

Based on the above facts, NMR can play a decisive role from in various stages of drug discovery i.e., from structural elucidation to identifying medicinally valid drug candidates. This article is an effort to throw a light on the basic principles involved in each and every technique of NMR, their advantages and limitations, along with its applicability.

Nuclear magnetic resonance (NMR) spectroscopy is the most widely applicable method for drug discovery and analysis. This technique provides a highly specific tool for identifying a drug substance containing impurities and residual solvents and their metabolites in biological media. It also provides a suitable analytical technique for their absolute quantification. In recent years, NMR spectroscopy has been increasingly used to monitor the cumulative drug release, drug dissolution, and diffusion coefficient of drugs from drug delivery systems in vitro and in vivo. Furthermore, this technique provides a better understanding of the release behaviors of drugs from drug delivery systems based on diffusion, dissolution, and osmosis mechanisms. Although early studies have been mainly qualitative in nature, these techniques can offer considerable information on release processes at the molecular level. Moreover, NMR spectroscopy has been used to detect structural changes that occur in drug delivery systems during the dissolution process. This review focuses on an overview of drug delivery systems and NMR spectroscopy and the application of NMR spectroscopy to drug release behaviors in drug delivery systems.

In the late 90’s, NMR based screening emerged as a powerful technique in the identification of new targeted molecule in drug discovery, at both industrial and academic levels. The capacity of finding ligands with low affinity has proved NMR to be a leader technique for Fragmented Based Drug Discovery (FBDD). This approach, complementary to HTS (High Throughput Screening), is based on the idea that it is easier to develop small and low affinity molecules endowed with BEI (Binding Efficiency Index) comparable to potent drug molecules with respect to the HTS identified molecules. Several NMR screening methods have been developed in the last 20 years to identify these fragments, and they were generally based on the observation of protein signals of interest (e.g. SAR by NMR) or on ligand signals (e.g. waterLOGSY). In this paper, different NMR techniques and their pharmaceutical applications will be summarized and discussed.

Nuclear magnetic resonance (NMR) utilizes spin changes at the nuclear level when radiofrequency energy is absorbed in the presence of magnetic field. Only nuclei with odd mass numbers (e.g., 1H, 13C, 15N) give NMR spectra because they have asymmetrical charge distribution and (2I+1) orientations. Since the discovery of NMR phenomenon in 1946 by Purcell and Bloch, it has been used for the study of both synthetic compounds and natural products. NMR has also been used to investigate dynamic molecular properties such as conformational isomerism, molecular asymmetry, hydrogen bonding, and keto-enol tautomerism. Analysis of structure-activity relationship with an NMR technique was introduced in 1996. Using this and other recently advanced NMR techniques, it is now possible to determine the binding site and pattern of a small molecule to its intended molecular target, an extremely powerful tool for the development of effective drugs. One of the most exciting new areas of research is the field of metabonomics, which relies on NMR spectroscopy of biofluids such as urine, plasma, and cerebrospinal fluid. In this review, we will focus on commonly used NMR-based screening approaches for cancer chemotherapeutics development. In doing so, we will first briefly introduce the theoretical aspect of the technique that lays the foundation of NMR-based methods. We will then discuss several different NMR techniques that are commonly used for studying interactions between model ligands and receptors. We will also describe examples of how the NMR spectroscopy has been applied to generate and optimize novel chemical classes of ligands to develop effective anticancer therapeutics. Finally, we will discuss about contributions of the metabonomic NMR spectroscopy to the analysis of drug metabolism and toxicity in the context of apoptosis and tumor control.

Recently, 1H, 31P, and 19F NMR magnetic resonance imaging MRI of radiosensitizers has been applied for measuring tumor and tissue oxygenation. In spite of the great importance of such bioreductive drugs, urthermore, surprisingly little detailed computational work on this subject has appeared. Thus, in this work a detailed computational study of BD is presented, calling special attention to the performance of various theoretical methods in reproducing the 13C and 15N, coupling-constant (H-N) data observed in solution. The most sophisticated approach involves density functional based Car–Parrinello molecular dynamics simulations (CPMD) in aqueous solution and averaging chemical shifts over snapshots from the trajectory. In the NMR calculations for these snapshots (performed at the B3LYP level), a small number of discrete water molecules are retained, and the remaining bulk solution effects are included via a polarizable continuum model (PCM). A similarly good accord with experiment is obtained from much less involved, static geometry optimization and NMR computation of pristine 1 employing a PCM approach. Solvent effects on chemical shift are not due to changes in geometric parameters upon solvation, but arise from the direct response of the electronic wave function to the presence of the solvent, which can be represented by discrete molecules and/or the dielectric bulk.

The application of NMR technology in drug development is generally adopted to describe the structure of a drug substance and its degradation products under standard stressed conditions. As far as drug products are concerned, standard stability conditions are recommended by international health authorities (ICH conditions), while the stability profile of a drug substance can be defined by non-standard protocols. Both proton and carbon NMR spectroscopic technologies are powerful methods for the determination of a drug substance chemical structure and related degradation products. Unconventional use of NMR in drug development is described in this section, namely NMR applications in the development of drugs containing cyclodextrin inclusion complexes. Cyclodextrins (CD) are cyclic oligosaccharides whose lipophilic cavity and hosting capacity make them ideal for forming inclusion complexes with lipophilic drugs. Generally, this complexation involves the inclusion of a ‘guest’ molecule into the cavity of a CD ‘host’ molecule, with no covalent bonding. Inclusion and/or molecular dispersion of a drug in cyclodextrin are made possible by the partitioning of the dissolved drug between the aqueous phase and the hydrophobic cyclodextrin cavity followed by specific molecular interactions, including hydrogen bonding and hydrophobic interactions between the drug and cyclodextrin. Proton NMR is used to confirm the host/guest inclusion by the chemical shift of the cyclodextrin internal protons, which are disturbed by the lipophilic guest. Examples of approved drug products containing cyclodextrins and two detailed case studies involving NMR applications in the development of water soluble diclofenac sodium and progesterone for parenteral use are reported in this article.

We present a concise review on current advances in the field of application of Nuclear Magnetic Resonance (NMR) in Structure-Activity Relationship (SAR) study of nanostructure drugs, particularly of paclitaxel-loaded nanocarriers. A short introduction of NMR to nonspecialists is also included. The review is separated into two parts, the first discusses in broader context the concepts and potentials of drug-loaded nanocarriers as the modern tumour active targeting systems and the second focuses on the paclitaxel-loaded nanocarriers, especially on conformations and tubulin-binding forms of paclitaxel. The application of nanoscience in pharmaceutical researches explodes very fast with many anti-tumour drugs recently been approved to the market. We survey only the important and latest issues in this field.