Seeking Nonspecific Binding: Assessing the Reliability of Tissue Dilutions for Calculating Fraction Unbound.

It has become commonplace (270+ article citations to date) to measure the fraction unbound (FrUn) of drugs in tissue homogenates and diluted plasma and then use a Correction Factor Equation (CFE) to extrapolate to the undiluted state. The CFE is based on assumptions of nonspecific binding with experimental use of very low drug concentrations. There are several possible determinants of apparent nonspecific binding as measured by methods such as equilibrium dialysis: true macromolecule binding and lipid partitioning along with receptor, enzyme, and transporter interactions. Theoretical calculations based on nonlinear protein binding indicate that the CFE will be most reliable to obtain FrUn when added drug concentration is small, binding constants are weak, protein concentrations are relatively high, and tissue dilution is minimal. When lipid partitioning is the sole factor determining apparent tissue binding, the CFE should be perfectly accurate. Use of very low drug concentrations, however, makes it more likely that specific binding to receptors and other targets may occur, and thus FrUn may reflect some binding to such components. Inclusion of trapped blood can clearly cause minor to marked discrepancies from purely tissue binding alone, which can be corrected. Furthermore, assessment of the occurrence of ionization/pH shifts, drug instability, and tissue metabolism may be necessary. Caution is needed in the use and interpretation of results from tissue dilution studies and other assessments of nonspecific binding, particularly for very strongly bound drugs with very small FrUn values and in tissues with metabolic enzymes, receptors, and trapped blood. SIGNIFICANCE STATEMENT: The use of tissue, plasma, and cell preparations to help obtain fraction unbound and tissue-to-plasma partition coefficients in pharmacokinetics has grown commonplace, especially for brain. This report examines theoretical, physiological, and experimental issues that need consideration before trusting such measurements and calculations.

Pharmacokinetics of the 1.5 mg levonorgestrel emergency contraceptive in women with normal, obese and extremely obese body mass index.

OBJECTIVE: To assess the pharmacokinetics (PK) of levonorgestrel after 1.5 mg oral doses (LNG-EC) in women with normal, obese and extremely obese body mass index (BMI). STUDY DESIGN: The 1.5 mg LNG dose was given to healthy, reproductive-age, ovulatory women with normal BMI (mean 22.0), obese (mean 34.4), and extremely obese (mean 46.6 kg/m2) BMI. Total serum LNG was measured over 0 to 96 h by radioimmunoassay while free and bioavailable LNG were calculated. The maximum concentration (Cmax), time to maximum concentration (Tmax), and area under the curve (AUC) of LNG were assessed. Pharmacokinetic parameters calculated included half-life (t1/2), clearance (CL) and volume of distribution (Vss). RESULTS: Ten normal-BMI, 11 obese-BMI, 5 extremely obese-BMI women were studied. After LNG-EC, mean total LNG metrics were lower in the obese and extremely obese groups compared to normal (Cmax 10.5 and 10.5 versus 16.2 ng/mL, both p<.01; AUC 208 and 197 versus 360 h × ng/mL, both p<.05). Mean bioavailable LNG Cmax was lower in obese (7.03 ng/mL, p<.05) and extremely obese (7.53 ng/ml, p=.198) compared to normal BMI (9.39 ng/mL). Mean bioavailable LNG AUC values were lower in obese and extremely obese compared to normal (131.6 and 127.5 vs 185.0 h × ng/mL, p<.05 for both). CONCLUSIONS: Obese and extremely obese women were exposed to lower total and bioavailable LNG than normal BMI women. IMPLICATIONS: Lower 'bioavailable' (free plus albumin bound) LNG AUC in obese women may play a role in the purported reduced efficacy of LNG-EC in obese users.

Pharmacokinetics and pharmacodynamics of liposomal chemophototherapy with short drug-light intervals.

Chemophototherapy (CPT) merges photodynamic therapy with chemotherapy and can substantially enhance drug delivery. Using a singular liposomal formulation for CPT, we describe a semi-mechanistic pharmacokinetic-pharmacodynamic (PK/PD) model to investigate observed antitumor effects. Long-circulating, sterically-stabilized liposomes loaded with doxorubicin (Dox) stably incorporate small amounts of a porphyrin-phospholipid (PoP) photosensitizer in the bilayer. These were administered intravenously to mice bearing low-passage, patient-derived pancreatic cancer xenografts (PDX). Dox PK was described with a two-compartment model and tumor drug disposition kinetics were modeled with first-order influx and efflux rates. Tumor irradiation with 665 nm laser light (200 J/cm2) 1 h after liposome administration increased tumor vascular permeabilization and drug accumulation, which was accounted for in the PK/PD model with increased tumor influx and efflux rates by approximately 12- and 4- fold, respectively. This modeling approach provided an overall 7-fold increase in Dox area under the curve in the tumor, matching experimental data (7.4-fold). A signal transduction model based on nonlinear direct cell killing accounted for observed tumor growth patterns. This PK/PD model adequately describes the CPT anti-PDX tumor response based on enhanced drug delivery at the short drug-light interval used.

Assessment of Three-Drug Combination Pharmacodynamic Interactions in Pancreatic Cancer Cells.

The pharmacodynamic interactions among trifluoperazine (TFP), gemcitabine (GEM), and paclitaxel (PTX) were assessed in pancreatic cancer cells (PANC-1). The phenothiazine TFP was chosen for its potential activity on cancer stem cells, while GEM and PTX cause apoptosis. Effects of each drug alone and in various combinations on cell growth inhibition of PANC-1 cells were studied in vitro to determine the drug-specific parameters and assess the nature of drug interactions. Joint inhibition (JI) and competitive inhibition (CI) equations were applied with a ψ interaction term. TFP fully inhibited growth of cells (Imax = 1) with an IC50 = 9887 nM. Near-maximum inhibition was achieved for GEM (Imax = 0.825) and PTX (Imax= 0.844) with an IC50 = 17.4 nM for GEM and IC50 = 7.08 nM for PTX. Estimates of an interaction term ψ revealed that the combination of TFP-GEM was apparently synergistic; close to additivity, the combination TFP-PTX was antagonistic. The interaction of GEM-PTX was additive, and TFP-GEM-PTX was synergistic but close to additive. The combination of TFP IC60-GEM IC60-PTX IC60 seemed optimal in producing inhibition of PANC-1 cells with an inhibitory effect of 82.1-90.2%. The addition of ψ terms to traditional interaction equations allows assessment of the degree of perturbation of assumed mechanisms.