A comprehensive regulatory and industry review of modeling and simulation practices in oncology clinical drug development.

Exposure-response (E-R) analyses are an integral component in the development of oncology products. Characterizing the relationship between drug exposure metrics and response allows the sponsor to use modeling and simulation to address both internal and external drug development questions (e.g., optimal dose, frequency of administration, dose adjustments for special populations). This white paper is the output of an industry-government collaboration among scientists with broad experience in E-R modeling as part of regulatory submissions. The goal of this white paper is to provide guidance on what the preferred methods for E-R analysis in oncology clinical drug development are and what metrics of exposure should be considered.

Correction to: Physiologically Based Pharmacokinetic Modeling of CFTR Modulation in People with Cystic Fibrosis Transitioning from Mono or Dual Regimens to Triple-Combination Elexacaftor/Tezacaftor/Ivacaftor.

The original version of this article unfortunately contained a mistake.

Physiologically Based Pharmacokinetic Modeling of CFTR Modulation in People with Cystic Fibrosis Transitioning from Mono or Dual Regimens to Triple-Combination Elexacaftor/Tezacaftor/Ivacaftor.

INTRODUCTION: The triple-combination (TC) cystic fibrosis transmembrane conductance regulator (CFTR) modulator regimen elexacaftor, tezacaftor, and ivacaftor was shown to be safe and efficacious in phase 3 trials of people with cystic fibrosis (pwCF) ≥ 12 years of age with ≥ 1 F508del-CFTR allele. Here, a simulation study predicted ivacaftor, tezacaftor, and elexacaftor exposures and impacts on CFTR modulation following transition from ivacaftor [a cytochrome P450 3A (CYP3A) substrate], lumacaftor (a CYP3A inducer)/ivacaftor, or tezacaftor/ivacaftor to TC. METHODS: Physiologically based pharmacokinetic (PBPK) modeling was used to evaluate plasma exposures during transition from mono- or dual-combination CFTR modulator regimens to TC. PBPK models were parameterized using data from human hepatocytes to account for CYP3A induction by lumacaftor and validated to match clinical data from healthy volunteers and pwCF. Using dosing regimens for pwCF ≥ 12 years of age, simulations were performed for ivacaftor, lumacaftor/ivacaftor, and tezacaftor/ivacaftor dosing for 14 days followed by immediate transition to elexacaftor/tezacaftor/ivacaftor dosing for 14 days. Drug exposures during transitions were compared with respective half-maximal effective concentrations (EC50) estimated from efficacy endpoint data from clinical studies. RESULTS: In simulations of immediate transition from ivacaftor or tezacaftor/ivacaftor to TC, the preceding treatment had no impact on ivacaftor, tezacaftor, or elexacaftor exposures. In simulations of immediate transition from lumacaftor/ivacaftor to TC, ivacaftor exposure decreased to 64% of maximum effective concentration (EC), due to reduction in ivacaftor dose and residual CYP3A4 induction, then returned to 90-95% of maximum EC. Lumacaftor-mediated CYP3A induction resolved within approximately 2 weeks. In all simulations, ivacaftor, tezacaftor, and elexacaftor exposures approached steady state within 2 weeks following transition and, at all times, ivacaftor and ≥ 1 CFTR corrector remained above EC50. CONCLUSION: PBPK modeling indicates that immediate transition to the elexacaftor/tezacaftor/ivacaftor regimen from an ivacaftor, lumacaftor/ivacaftor, or tezacaftor/ivacaftor regimen results in sustained CFTR modulation in pwCF ≥ 12 years of age.

Assessment of Transporter-Mediated and Passive Hepatic Uptake Clearance Using Rifamycin-SV as a Pan-Inhibitor of Active Uptake.

The use of in vitro data for the quantitative prediction of transporter-mediated clearance is critical. Central to this evaluation is the use of hepatocytes, since they contain the full complement of transporters and metabolic enzymes. In general, uptake clearance (CLuptake) is evaluated by measuring the appearance of compound in the cell. Passive clearance (CLpd) is often determined by conducting parallel studies at 4 °C or by attempting to saturate uptake pathways. Both approaches have their limitations. Recent studies have proposed the use of Rifamycin-SV (RFV) as a pan-inhibitor of hepatic uptake pathways. In our studies, we confirm that transport activity of all major hepatic uptake transporters is inhibited significantly by RFV at 1 mM (OATP1B1, 1B3, and 2B1 = NTCP (80%), OCT1 (65%), OAT2 (60%)). Under these incubation conditions, we found that the free intracellular concentration of RFV is ∼175 µM and that several major CYPs and UGTs can be reversibly inhibited. Using this approach, we also determined CLuptake and CLpd of nine known OATP substrates across three different lots of human hepatocytes. The scaling factors generated for these compounds at 37 °C with RFV and 4 °C were found to be similar. The CLpd of passively permeable compounds like metoprolol and semagacestat were found to be higher at 37 °C compared to 4 °C, indicating a temperature effect on these compounds. In addition, our data also suggests that incorporation of medium concentrations into CLuptake and CLpd calculations may be critical for highly protein bound and highly lipophilic drugs. Overall, our data indicate that RFV, instead of 4 °C, can be reliably used to measure CLuptake and CLpd of drugs.

Physiologically Based Pharmacokinetic Model Qualification and Reporting Procedures for Regulatory Submissions: A Consortium Perspective.

This work provides a perspective on the qualification and verification of physiologically based pharmacokinetic (PBPK) platforms/models intended for regulatory submission based on the collective experience of the Simcyp Consortium members. Examples of regulatory submission of PBPK analyses across various intended applications are presented and discussed. European Medicines Agency (EMA) and US Food and Drug Administration (FDA) recent draft guidelines regarding PBPK analyses and reporting are encouraging, and to advance the use and acceptability of PBPK analyses, more clarity and flexibility are warranted.

Enhanced active extracellular polysaccharide production from Ganoderma formosanum using computational modeling.

Extracellular polysaccharide (EPS) is one of the major bioactive ingredients contributing to the health benefits of Ganoderma spp. In this study, response surface methodology was applied to determine the optimal culture conditions for EPS production of Ganoderma formosanum. The optimum medium composition was found to be at initial pH 5.3, 49.2 g/L of glucose, and 4.9 g/L of yeast extract by implementing a three-factor-three-level Box-Behnken design. Under this condition, the predicted yield of EPS was up to 830.2 mg/L, which was 1.4-fold higher than the one from basic medium (604.5 mg/L). Furthermore, validating the experimental value of EPS production depicted a high correlation (100.4%) with the computational prediction response model. In addition, the percentage of ß-glucan, a well-recognized bioactive polysaccharide, in EPS was 53±5.5%, which was higher than that from Ganoderma lucidum in a previous study. Moreover, results of monosaccharide composition analysis indicated that glucose was the major component of G. formosanum EPS, supporting a high ß-glucan percentage in EPS. Taken together, this is the first study to investigate the influence of medium composition for G. formosanum EPS production as well as its ß-glucan composition.

Extract of Ganoderma formosanum Mycelium as a Highly Potent Tyrosinase Inhibitor.

In this study, the inhibitory effect of Ganoderma formosanum mycelium extracts on tyrosinase, the central regulatory enzyme being responsible for cutaneous pigmentation, was investigated in both cell-free and cellular enzymatic systems, as well as in phenotype-based zebrafish model. Bioassay-guided purification indicated that the ethyl acetate fraction of G. fromosanum mycelium ethanolic extract (GFE-EA) demonstrated the highest inhibition toward cell-free tyrosinase (IC50 = 118.26 ± 13.34 ppm). The secreted and intracellular melanin of B16-F10 cells were reduced by GFE-EA through suppression of tyrosinase activity (IC50 = 102.27 ± 9.49 ppm) and its protein expression. Moreover, GFE-EA decreased surface pigmentation level of zebrafish via down-regulation of tyrosinase activity. Most of all, there is no significant difference in morphology and mortality between control and GFE-EA treated groups. Not only does GFE-EA exhibit similar depigmenting efficacy to kojic acid with lower dosage (approximately one-seventh of dose), but show less toxicity to zebrafish. It is worth noting that GFE-EA is extracted from mycelium, which subverts the general concept that mycelium lacks certain bioactivities possessed by fruit bodies. Altogether, it would appear that GFE-EA has great potential for application in the cosmetics industry.

The effect of ABCG2 and ABCC4 on the pharmacokinetics of methotrexate in the brain.

Methotrexate (MTX) is the cornerstone of chemotherapy for primary central nervous system lymphoma, yet how the blood-brain barrier (BBB) efflux transporters ABCG2 and ABCC4 influence the required high-dose therapy is unknown. To evaluate their role, we used four mouse strains, C57BL/6 (wild-type; WT), Abcg2(-/-), Abcc4(-/-), and Abcg2(-/-);Abcc4(-/-) (double knockout; DKO) to conduct brain microdialysis studies after single intravenous MTX doses of 50 mg/kg. When the area under the concentration-time curve for plasma (AUC(plasma)) was used to assess systemic exposure to MTX, the rank order was Abcc4(-/-) < WT < Abcg2(-/-) < Abcg2(-/-)Abcc4(-/-). Only the DKO exposure was significantly higher than that of the WT group (P < 0.01), a reflection of the role of Abcg2 in biliary excretion and Abcc4 in renal excretion. MTX brain interstitial fluid concentrations obtained by microdialysis were used to calculate the area under the concentration-time curve for the brain (AUC(brain)), which found the rank order of exposure to be WT < Abcc4(-/-) < Abcg2(-/-) < Abcg2(-/-)Abcc4(-/-) with the largest difference being 4-fold: 286.13 ± 130 µg*min/ml (DKO) versus 66.85 ± 26 (WT). Because the transporters affected the systemic disposition of MTX, particularly in the DKO group, the ratio of the AUC(brain)/AUC(plasma) or the brain/plasma partition coefficient Kp was calculated, revealing that the DKO strain had a significantly higher value (0.23 ± 0.09) than the WT strain (0.11 ± 0.05). Both Abcg2 and Abcc4 limited BBB penetration of MTX; however, only when both drug efflux pumps were negated did the brain accumulation of MTX significantly increase. These findings indicate a contributory role of both ABCG2 and ABCC4 to limiting MTX distribution in patients.

Impact of intestinal PepT1 on the kinetics and dynamics of N-formyl-methionyl-leucyl-phenylalanine, a bacterially-produced chemotactic peptide.

The primary purpose of this study was to evaluate the intestinal permeability (P(eff)) of N-formyl-methionyl-leucyl-phenylalanine (fMet-Leu-Phe), a bacterially derived chemotactic tripeptide, in the duodenum, jejunum, ileum, and colon of wild-type and PepT1 knockout mice. A secondary purpose was to determine if the presence of intestinal PepT1 translated into fMet-Leu-Phe directed neutrophil migration in these animals. Using an in situ single pass perfusion technique, the P(eff) of [(3)H]fMet-Leu-Phe was substantially reduced in the duodenum, jejunum, and ileum of PepT1 knockout mice as compared to wild-type animals. In contrast, the P(eff) of [(3)H]fMet-Leu-Phe in colon was unchanged between genotypes and about 5% of that in small intestine. Jejunal uptake of [(3)H]fMet-Leu-Phe was specific for PepT1 and saturable with an intrinsic K(0.5) of 1.6 mM. The peptide/histidine transporters PhT1 and PhT2 were not involved in [(3)H]fMet-Leu-Phe uptake. Myeloperoxidase activity (a measure of neutrophil migration) was significantly increased following 4 h perfusions of 10 µM fMet-Leu-Phe in the jejunum of wild-type mice and was abolished by 50 mM glycylglycine; no change was observed in the jejunum of PepT1 knockout mice. Likewise, fMet-Leu-Phe perfusions had no effect on myeloperoxidase activity in the colon of either genotype. In conclusion, these findings demonstrated that PepT1 had a major influence on the permeability of fMet-Leu-Phe in duodenum, jejunum, and ileum in wild-type mice and on inflammatory response in intestinal regions that expressed PepT1.

Significance and regional dependency of peptide transporter (PEPT) 1 in the intestinal permeability of glycylsarcosine: in situ single-pass perfusion studies in wild-type and Pept1 knockout mice.

The purpose of this study was to evaluate the role, relevance, and regional dependence of peptide transporter (PEPT) 1 expression and function in mouse intestines using the model dipeptide glycylsarcosine (GlySar). After isolating specific intestinal segments, in situ single-pass perfusions were performed in wild-type and Pept1 knockout mice. The permeability of [(3)H]GlySar was measured as a function of perfusate pH, dipeptide concentration, potential inhibitors, and intestinal segment, along with PEPT1 mRNA and protein. We found the permeability of GlySar to be saturable (K(m) = 5.7 mM), pH-dependent (maximal value at pH 5.5), and specific for PEPT1; other peptide transporters, such as PHT1 and PHT2, were not involved, as judged by the lack of GlySar inhibition by excess concentrations of histidine. GlySar permeabilities were comparable in the duodenum and jejunum of wild-type mice but were much larger than that in ileum (approximately 2-fold). A PEPT1-mediated permeability was not observed for GlySar in the colon of wild-type mice (<10% residual uptake compared to proximal small intestine). Moreover, GlySar permeabilities were very low and not different in the duodenum, jejunum, ileum, and colon of Pept1 knockout mice. Functional activity of intestinal PEPT1 was confirmed by real-time polymerase chain reaction and immunoblot analyses. Our findings suggest that a loss of PEPT1 activity (e.g., due to polymorphisms, disease, or drug interactions) should have a major effect in reducing the intestinal absorption of di-/tripeptides, peptidomimetics, and peptide-like drugs.

Interaction between anticonvulsants and human placental carnitine transporter.

PURPOSE: To examine the inhibitory effect of anticonvulsants (AEDs) on carnitine transport by the human placental carnitine transporter. METHODS: Uptake of radiolabeled carnitine by human placental brush-border membrane vesicles was measured in the absence and presence of tiagabine (TGB), vigabatrin (VGB), gabapentin (GBP), lamotrigine (LTG), topiramate (TPM), valproic acid (VPA), and phenytoin (PHT). The mechanism of the inhibitory action of TGB was determined. RESULTS: Most of the AEDs inhibited placental carnitine transport. Kinetic analyses showed that TGB had the greatest inhibitory effect [50% inhibitory concentration (IC50, 190 microM)], and the order of inhibitory potency was TGB > PHT > GBP > VPA > VGB, TPM > LTG. Further studies showed that TGB competitively inhibited carnitine uptake by the human placental carnitine transporter, suggesting that it may be a substrate for this carrier. CONCLUSIONS: Although the involvement of carnitine deficiency in fetal anticonvulsant syndrome requires further evaluation, potential interference with placental carnitine transport by several AEDs was demonstrated. Despite the higher inhibitory potency of TGB, given the therapeutic unbound concentrations, the results for VPA and PHT are probably more clinically significant.