Drug PAMPA parallel artificial membrane

The evaluation of the apparent ionization constants (i) can indicate in partition experiments the extent to which a charged form of the drug partitions into the octanol or liposome bilayer domains, (ii) can indicate in solubility measurements, the presence of aggregates in saturated solutions and whether the aggregates are ionized or neutral and the extent to which salts of dmgs form, and (iii) can indicate in permeability measurements, whether the aqueous boundary layer adjacent to the membrane barrier, Umits the transport of drugs across artificial phospholipid membranes [parallel artificial membrane permeation assay (PAMPA)] or across monolayers of cultured cells [Caco-2, Madin-Darby canine kidney (MDCK), etc.]. 

This book is written for the practicing pharmaceutical scientist involved in absorption-distribution-metabolism-excretion (ADME) measurements who needs to communicate with medicinal chemists persuasively, so that newly synthesized molecules will be more drug-like. ADME is all about a day in the life of a drug molecule (absorption, distribution, metabolism, and excretion). Specifically, this book attempts to describe the state of the art in measurement of ionization constants (p Ka), oil-water partition coefficients (log PI log D), solubility, and permeability (artificial phospholipid membrane barriers). Permeability is covered in considerable detail, based on a newly developed methodology known as parallel artificial membrane permeability assay (PAMPA). 

There are several approaches to estimating absorption using in vitro methods, notably Caco-2 and MDCK cell-based methods or using methods that assess passive permeability, for example the parallel artificial membrane permeation assay (PAMPA) method. These are reviewed elsewhere in this book. The assays are very useful, and usually have an important role in the screening cascades for drug discovery projects. However, as discussed below, the cell-based assays are not without their drawbacks, and it is often appropriate to use ex vivo and/or in vivo absorption assays. 

Table 18.3 Parallel artificial membrane permeability assay (PAMPA) permeability and human absorption for 19 drug compounds.

Solubility and permeability were measured by a high throughput solubility assay and parallel artificial membrane permeation assay (PAMPA), respectively [56], The assays categorized 14 out of 18 drugs based on the BCS consistent with their known solubility and permeability characteristics [56],... 

HTS plates permit to determine drug permeability across a cell monolayer with a throughput similar to that of the parallel artificial membrane permeation assay (PAMPA), which measures rate of diffusion across a lipid layer.46 As is the case with PAMPA, the tiny surface area of the filters of the 96-well HTS presents an analytical challenge for compounds with low-to-moderate permeability. 

One decade has passed since the parallel artificial membrane permeation assay (PAM PA) was first introduced in 1998 [47]. Since then, PAM PA rapidly gained wide popularity in drug discovery [3, 48-51]. Today, PAMPA is the most widely used physicochemical membrane permeation model. The term PAMPA is nowusedas the general name for a plate-based (HTS enabled), biter-supported (filter immobilized) artificial membrane. Typically, phospholipids dissolved in an organic solvent are impregnated into the filter to construct a PAMPA membrane. 

Kansy et al. (1998) proposed the use a parallel artificial membrane permeation assay (PAMPA) as a high-throughput alternative to Caco-2 monolayers for the prediction of passive drug permeation. In the PAMPA approach, aqueous... 

Van Dijck A, Masungi C, MenschJ, Borremans C, Willems B, Mackie C, Brewster M, and Noppe, M. Parallel Artificial Membrane Permeability Assay (PAMPA) Combined with a 10-Day Multiscreen CACO-2 Cell Culture for Prediction of Passive and Active Absorption of Drugs. 2003 American Association of Pharmaceutical Scientists Annual Meeting and Exposition, Salt Lake City, Utah, USA, October 26-30, 2003. 

The prediction of the important structural features that affect intestinal permeability is useful information to obtain early in the drug discovery process. The two most common models used to obtain fast, high-throughput measurements are the parallel artificial membrane permeation assay (PAMPA) and the cell line assays that feature cultured human colon adenocarcinoma cells (Caco-2). Each method uses a surrogate model to mimic intestinal absorption followed by LC-MS analysis. 

Figure 9.8 Parallel artificial membrane permeability assay Papp values versus literature values, for human oral absorption, for a set of 93 drugs. Note circled areas, where PAMPA data were not predictive, including 45, griseofulvin, for which bioavailability is known to be very formulation-dependent. (Reprinted with permission from Zhu, C., et al. A comparative study of artificial membrane permeability assay for high throughput profiling of drug absorption potential. Eur. J. Med. Chem. 2002, 37, 399 07, copyright 2002, Elsevier).

The complementarity between PAMPA and Caco-2 measurements and a similar staging scheme for their use in drug discovery is advocated in Kerns, E.Fl., et al. Combined application of parallel artificial membrane permeability assay and Caco-2 permeability assays in drug discovery. J. Pharm. Sci. 2004, 93, 1440-1453. 

Artificial Systems (PAMPA) Some artificial systems are widely used as an alternative to Caco-2 cell culture to evaluate drug permeability. An example of these is the parallel artificial membrane permeation assay (PAMPA). PAMPA is usually performed in a 96-well microtiter plate format that consists of test wells fixed with an artificial hydrophobic membrane and reference wells (Kansy et al., 1998). Test compounds are applied at one side of the membrane, and the permeability rates are monitored at the receiving side. The properties of the membrane (lipophilicity and thickness) control the rate of permeability. Different membranes could be designed to mimic the passive... 

Masungi C, Mensch J, Van Dijck A, Borremans C, Willems B, Mackie C, Noppe M, Brewster ME. Parallel artificial membrane permeability assay (PAMPA) combined with a 10-day multiscreen Caco-2 cell culture as a tool for assessing new drug candidates. Pharmazie 2008 63(3) 194-199. 

Permeability and Associated Assays Orally administered drugs need to cross the intestinal epithelium cell layer to reach the systemic circulation. Therefore, membrane permeability is a major determining factor of intestinal absorption and oral bioavailability for drug molecules. Several in vitro systems mimicking the epithelium cell layer are routinely used in ADME profiling to assess membrane permeability of NCEs. The parallel artificial membrane permeability assay (PAMPA) uses a dual chamber sandwich plate separated by an artificial lipid membrane to simulate the epithelium layer (Figure 6.6A) [75]. Compounds are... 

From a methodological point of view, tite PAMPA-BBB system is quite simple. The lipids, dissolved in dodecane, are soaked with a filter mounted in a two-compartment chamber. The drug is added to the donor compartment (which can be either the upper or lower chamber), and its passage through the artificial membrane is measured in the acceptor compartment (Fig. 14.17). A standard compound with well-characterized permeability proper-hes (e.g., verapamil) is tested in parallel. Compounds that readily cross the blood-brain barrier have an in vitro permeability (P,) > 2.7 10" cm s in the PAMPA assay. On the opposite, drugs with low blood-brain barrier permeation have a < 0.710 cm s". Beside... 

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