scholarly journals Virtual experiments provide plausible explanations for IVIVE underpredictions of hepatic clearance

2020 ◽  
Author(s):  
Preethi Krishnan ◽  
Andrew K. Smith ◽  
Glen E.P. Ropella ◽  
Lopamudra Dutta ◽  
Ryan C. Kennedy ◽  
...  
Keyword(s):  
Discrete Event ◽  
Hepatic Clearance ◽  
Scaling Factor ◽  
Cell Entry ◽  
Software Systems ◽  
Removal Rates ◽  
Unbound Fraction ◽  
Virtual Experiments ◽  
Hepatocyte Cultures ◽  
Rat Livers

AbstractWe demonstrate how results of virtual experiments can suggest mechanism-based explanations for IVIVE underpredictions of hepatic clearance. We use agent-oriented, discrete-event methods. We experiment on software analogies of rats and hepatocyte cultures dosed identically with objects that mimic two idealized compounds, vC1 (not cleared) and vC2 (removal maximized). Hepatocytes in cultures and rat livers interact identically with vC1 and vC2. The rules governing vC1 and vC2 movements and interactions with hepatocytes are the same in both systems. The two software systems are independent; to be useful in discovering plausible explanations of IVIVE inaccuracies, we require that, in the absence of hepatocyte exposure differences, the culture-to-rat scaling factor = 1.0 for all corresponding measures. The probability of cell entry (pEnter) maps directly to the unbound-fraction of drug used by IVIVE methods. For vC1 (vC2), we achieve that validation target for pEnter = 0.05-1.0 (1.0). However, for pEnter = 0.05-0.8, vC2 removal rates during culture experiments underpredicted corresponding removal rates in rats. The magnitude of the underpredictions increases with decreasing pEnter. For example, using pEnter = 0.1 (0.3), peak vC2 removal rates in culture experiments underpredict corresponding removal rates in rats by 3.2 (1.4) fold. Underpredictions are a consequence of the biomimetic periportal-to-pericentral organization of hepatocytes within virtual livers are absent in virtual cultures. In rats, pericentral hepatocytes do more of the vC2 removal work. The results suggest that using IVIVE methods that abstract away influential features of hepatocyte organization within livers may contribute to IVIVE underpredictions.Significance StatementFrom experiments on virtual (software) counterparts of rats and hepatocyte cultures, we learned that IVIVE underpredictions are a consequence of the biomimetic periportal-to-pericentral organization of hepatocytes within virtual livers being absent in virtual cultures. Using IVIVE methods that abstract away influential features of hepatocyte organization within livers may contribute in part to some IVIVE underpredictions.Visual Abstract

scholarly journals Hepatocyte Organization Affects the Translation of Clearance from In Vitro to In Vivo

10.29007/t4kv ◽  
2020 ◽  
Author(s):  
Lopamudra Dutta ◽  
Preethi Krishnan ◽  
Andrew Smith ◽  
Ryan Kennedy ◽  
Glen Ropella ◽  
...  
Keyword(s):  
Hepatic Clearance ◽  
Intrinsic Clearance ◽  
Quasi Steady State ◽  
Exposure Rate ◽  
Virtual Experiments ◽  
Mitigating Factors ◽  
Virtual Mouse ◽  
In Vivo Extrapolation

An improved understanding of in vivo ⇔ in vitro changes is crucial in identifying and mitigating factors contributing to in vitro–in vivo extrapolation (IVIVE) inaccuracies in predicting the hepatic clearance of substances. We argue that a model mechanism-based virtual culture (vCulture) ⇔ virtual mouse (vMouse) (or vRat or vHuman) experiment approach can identify factors contributing to IVIVE disconnects. Doing so depends on having evidence that six Translational Requirements have been achieved. We cite evidence that the first four have been achieved. The fifth Requirement is that differences in measures of vCompound disposition between vCulture and vMouse are attributable solely to the micro-architectural, physiomimetic features, and uncertainties built into the vLiver and vMouse but are absent from the vCulture. The objective of this work is to first improve on a vCulture architecture used previously and then use results of virtual experiments to verify that its use enables the fifth Translational Requirement to be achieved. We employ two different idealized vCompounds, which map to highly permeable real compounds at the extreme ends of the intrinsic clearance spectrum. Virtual intrinsic clearance = Exposure rate per vHPC. At quasi-steady state, results for vCompound-1 are independent of the dosing rate. The average per-vHPC Exposure rates (taken over the whole vLiver in vMouse experiments) are the same (within the variance of the Experiments) as those in vCulture. However, they are location dependent within the vLiver. For vCompound-2, there are dosing rate differences and average per-vHPC Exposure rates within the vLiver are also location dependent. When we account for dosing rate differences, we see again that average per-vHPC Exposure rates averaged over the whole vLiver in vMouse experiments are the same as those in vCulture. Thus, the differences in per vHPC Exposure rate within the vLiver for both vCompounds are attributable solely to the micro-architectural and physiomimetic features built into the vLiver and vMouse but are absent from the vCulture. The results verify that the fifth Translational Requirement has been achieved.

Metabolism of intact parathyroid hormone in isolated perfused rat liver and kidney

1988 ◽  
Vol 254 (6) ◽  
pp. E740-E748 ◽  
Author(s):  
H. Daugaard ◽  
M. Egfjord ◽  
K. Olgaard
Keyword(s):  
Parathyroid Hormone ◽  
High Performance ◽  
Hepatic Clearance ◽  
Isolated Perfused Rat Liver ◽  
Intact Parathyroid Hormone ◽  
Human Parathyroid Hormone ◽  
Intact Pth ◽  
Glandular Secretion ◽  
Liver And Kidney ◽  
Rat Livers

Metabolism of synthetic human parathyroid hormone (PTH) 2 X 10(-10) to 5 X 10(-9) M was studied in 16 isolated perfused rat kidneys and 12 isolated perfused rat livers. Organ clearances were measured by assays specific for intact PTH. Production of fragments was analyzed by high-performance liquid chromatography (HPLC) and radioimmunoassays specific for NH2-terminal, midmolecule, and COOH-terminal PTH. The livers cleared intact PTH and NH2-terminal immunoreactive PTH (iPTH) at the same rate. Midmolecule iPTH was cleared significantly (P less than 0.001) slower, as was COOH-terminal iPTH (P less than 0.005), and HPLC studies demonstrated production of midmolecule/COOH-terminal PTH fragments, while no NH2-terminal fragments were found. Clearance in the kidneys of intact PTH and of NH2-terminal, midmolecule, and COOH-terminal iPTH was not significantly different from clearance of inulin. No clearance of intact PTH was found in nonfiltering kidneys. HPLC studies did not demonstrate release of any PTH fragments from the kidneys. In conclusion, the liver was not selective for intact PTH, and differential hepatic clearance, possibly together with direct glandular secretion, may contribute to the predominance of COOH-terminal PTH fragments in plasma.

scholarly journals Exposure Measurements on Biomimetic Lobules Using Virtual Experiments to Help Improve IVIVE

10.29007/hjfc ◽  
2020 ◽  
Author(s):  
Preethi Krishnan ◽  
Lopamudra Dutta ◽  
Andrew Smith ◽  
Glen Ropella ◽  
Ryan Kennedy ◽  
...  
Keyword(s):  
Hepatic Clearance ◽  
Intrinsic Clearance ◽  
Central Vein ◽  
Unbound Fraction ◽  
Hepatic Disposition ◽  
Liver Model ◽  
New Chemical Entities ◽  
Model Drug

The in vitro-in vivo extrapolation (IVIVE) methods used currently to predict the hepatic clearance of new chemical entities are plagued by poorly understood inaccuracies. To begin identifying plausible sources, we challenge two of core hypotheses. Hypothesis-1: the intralobular micro-anatomical organization of hepatocytes (HPCs) can be abstracted away. By accepting that hypothesis, one can assume that intrinsic clearance per HPC is essentially the same in vitro and in vivo, and thus an IVIVE method can employ a simplified liver model, typically the “well-stirred” liver model. Hypothesis-2: when the simplified liver model is the “parallel tube model,” drug concentration decreases exponentially from portal to central vein. When either simplified liver model is used, a core assumption is that intrinsic clearance is directly proportional to the unbound fraction of drug. A barrier to progress has been the fact that it is currently infeasible to challenge the two hypotheses using wet-lab experiments. In this work, we challenge virtual counterparts of the two hypotheses by experimenting on virtual mice in which hepatic disposition and clearance are consequences of concretized model mechanisms that have met several demanding requirements, including the following. The virtual liver’s structure and organization are strongly analogous to those of an actual liver, and the hepatic disposition and clearance of several virtual compounds have achieved quantitative validation targets. We study two virtual compounds. Compound-1 simulates the extreme of low-clearance, highly permeable compounds. Compound-2 simulates a highly permeable compound exhibiting maximum intrinsic clearance. We simulate changes in unbound fraction by changing the probability (pEnter) that a Compound-1 or -2 will enter an adjacent HPC during a simulation cycle. Compound-1 and -2 HPC exposure rates do not decrease from portal to central vein: they increase, and that contradicts both hypotheses. Further, the relationship between exposure rates and pEnter is nonlinear. The insights achieved help explain the frequently reported underprediction of in vivo hepatic clearance values. We suggest that IVIVE methods can be improved by utilizing a liver model that couples a biomimetic representation of intralobular HPC organization with biomimetic representations of intrahepatic disposition dynamics.

scholarly journals Assessing Toxicokinetic Uncertainty and Variability in Risk Prioritization

2019 ◽  
Vol 172 (2) ◽  
pp. 235-251 ◽  
Author(s):  
John F Wambaugh ◽  
Barbara A Wetmore ◽  
Caroline L Ring ◽  
Chantel I Nicolas ◽  
Robert G Pearce ◽  
...  
Keyword(s):  
Measurement Uncertainty ◽  
Uncertainty Propagation ◽  
Hepatic Clearance ◽  
Biological Variability ◽  
Vitro Assay ◽  
Unbound Fraction ◽  
Risk Prioritization ◽  
Uncertainty Estimates

Abstract High(er) throughput toxicokinetics (HTTK) encompasses in vitro measures of key determinants of chemical toxicokinetics and reverse dosimetry approaches for in vitro-in vivo extrapolation (IVIVE). With HTTK, the bioactivity identified by any in vitro assay can be converted to human equivalent doses and compared with chemical intake estimates. Biological variability in HTTK has been previously considered, but the relative impact of measurement uncertainty has not. Bayesian methods were developed to provide chemical-specific uncertainty estimates for 2 in vitro toxicokinetic parameters: unbound fraction in plasma (fup) and intrinsic hepatic clearance (Clint). New experimental measurements of fup and Clint are reported for 418 and 467 chemicals, respectively. These data raise the HTTK chemical coverage of the ToxCast Phase I and II libraries to 57%. Although the standard protocol for Clint was followed, a revised protocol for fup measured unbound chemical at 10%, 30%, and 100% of physiologic plasma protein concentrations, allowing estimation of protein binding affinity. This protocol reduced the occurrence of chemicals with fup too low to measure from 44% to 9.1%. Uncertainty in fup was also reduced, with the median coefficient of variation dropping from 0.4 to 0.1. Monte Carlo simulation was used to propagate both measurement uncertainty and biological variability into IVIVE. The uncertainty propagation techniques used here also allow incorporation of other sources of uncertainty such as in silico predictors of HTTK parameters. These methods have the potential to inform risk-based prioritization based on the relationship between in vitro bioactivities and exposures.

Ultrastructural changes in isolated rat liver perfused with cyclic heptapeptide toxins from norwegian freshwater cyanobacteria (blue-green algae)

1987 ◽  
Vol 45 ◽  
pp. 858-859
Author(s):  
A.S. Dabholkar ◽  
W.W. Carmichael ◽  
K. Berg ◽  
J. Wyman
Keyword(s):  
Ultrastructural Changes ◽  
Sprague Dawley ◽  
Isolated Rat Liver ◽  
Blue Green Algae ◽  
Sprague Dawley Rats ◽  
Cyclic Heptapeptide ◽  
Intracellular Changes ◽  
Oscillatoria Agardhii ◽  
Rat Livers ◽  
Isolated Rat

Intracellular changes in the hepatocytes of isolated rat livers perfused with cyclic heptapeptide toxins are described. The toxins used are 1) -Ala-Leu- β-methyl isoAsp-Arg-ADDA-isoGlu-mdha (M.W. 944) from Microcystis aeruginosa- Lake Akersvatn, Norway; 2) -Ala-Arg-isoAsp-Arg-ADDA-isoGlu-mdha (M.W. 1023) from Oscillatoria agardhii var. - Lake Kolbatnvatn, Norway; 3) -Ala-Arg-isoAsp-Arg-ADDA-isoGlu-dha (M.W. 1009) from Oscillatoria agardhii var. isothrix - Lake Froylandsvatn, Norway. Approximate LD intraperitoneal mouse for the toxins is 50, 500 and 1000 μg/kg respectively.Livers were removed from male Sprague Dawley rats and perfused for 15 min with a blood-free perfusate (50 ml) followed by 60 min with perfusate containing i) 25, 50, or 200 μg of M. aeruginosa toxin ii) 50, 250, 500 or 1000 μg of O. agardhii var. toxin and iii) 1000, 2000, 2500 or 5000 μg of O. agardhii var. isothrix toxin. Control livers were perfused for 75 min with the blood-free perfusate.

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