The development of antibody-drug conjugates (ADCs) has significantly been advanced in the past decade given the improvement of payloads, linkers and conjugation methods. In particular, linker design ...plays a critical role in modulating ADC stability in the systemic circulation and payload release efficiency in the tumors, which thus affects ADC pharmacokinetic (PK), efficacy and toxicity profiles. Previously, we have investigated key linker parameters such as conjugation chemistry (e.g., maleimide vs. disulfide), linker length and linker steric hindrance and their impacts on PK and efficacy profiles. Herein, we discuss our perspectives on development of integrated strategies for linker design to achieve a balance between ADC stability and payload release efficiency for desired efficacy in antigen-expressing xenograft models. The strategies have been successfully applied to the design of site-specific THIOMAB
TM
antibody-drug conjugates (TDCs) with different payloads. We also propose to conduct dose fractionation studies to gain guidance for optimal dosing regimens of ADCs in pre-clinical models.
Antibody-drug conjugates (ADC) are designed to selectively bind to tumor antigens via the antibody and release their cytotoxic payload upon internalization. Controllable payload release through ...judicious design of the linker has been an early technological milestone. Here, we examine the effect of the protease-cleavable valine-citrulline VC(S) linker on ADC efficacy. The VC(S) linker was designed to be cleaved by cathepsin B, a lysosomal cysteine protease. Surprisingly, suppression of cathepsin B expression via CRISPR-Cas9 gene deletion or shRNA knockdown had no effect on the efficacy of ADCs with VC(S) linkers armed with a monomethyl auristatin E (MMAE) payload. Mass spectrometry studies of payload release suggested that other cysteine cathepsins can cleave the VC(S) linker. Also, ADCs with a nonprotease-cleavable enantiomer, the VC(R) isomer, mediated effective cell killing with a cysteine-VC(R)-MMAE catabolite generated by lysosomal catabolism. Based on these observations, we altered the payload to a pyrrolo2,1-c1,4benzodiazepine dimer (PBD) conjugate that requires linker cleavage in order to bind its DNA target. Unlike the VC-MMAE ADCs, the VC(S)-PBD ADC is at least 20-fold more cytotoxic than the VC(R)-PBD ADC. Our findings reveal that the VC(S) linker has multiple paths to produce active catabolites and that antibody and intracellular targets are more critical to ADC efficacy. These results suggest that protease-cleavable linkers are unlikely to increase the therapeutic index of ADCs and that resistance based on linker processing is improbable.
.
Drug discovery and development involve the utilization of in vitro and in vivo experimental models. Different models, ranging from test tube experiments to cell cultures, animals, healthy human ...subjects, and even small numbers of patients that are involved in clinical trials, are used at different stages of drug discovery and development for determination of efficacy and safety. The proper selection and applications of correct models, as well as appropriate data interpretation, are critically important in decision making and successful advancement of drug candidates. In this review, we discuss strategies in the applications of both in vitro and in vivo experimental models of drug metabolism and disposition.
Drug metabolism is critical to many aspects of drug safety and efficacy. Thus, understanding the various intricacies of drug metabolism helps to avoid possible pitfalls and contributes to drug design ...during the discovery stage and to drug characterization during the development stage. Drug metabolism expertise was mainly taught at the universities and prominent professors have contributed to the field and here, we like to acknowledge Professor Thomas Baillie contribution to the field. Over the years, we have deployed our learnings in both drug discovery and development at Genentech. Here, we describe case studies from novel reaction mechanisms to novel types of metabolites. Taken together, we describe our lessons in a manner that allows the reader to understand the significance of the lessons and the thinking behind the experimental design. Here, we describe 10 case studies that highlight our experience in the past decade with the first 5 case studies capturing non-cytochrome P450 enzymatic reactions such as the discovery of a novel hydrolytic reaction by aldehyde oxidase (Case Study 1). The remaining 5 case studies describe cytochrome P450-derived reactions including various types of cytochrome P450-mediated reactions that resulted in ring formation or cleavage (Case Studies 6–9). The last case study (Case Study 10), emphasizes the importance of exploring ways to understand binding and enzyme kinetics, and explores co-incubation of 1-aminobenzotriazole (ABT) with warfarin in CYP2C9 metabolism.
The metabolism and disposition of (14)Capixaban, an orally bioavailable, highly selective, and direct acting/reversible factor Xa inhibitor, was investigated in 10 healthy male subjects without ...(group 1, n=6) and with bile collection (group 2, n=4) after a single 20-mg oral dose. Urine, blood, and feces samples were collected from all subjects. Bile samples were also collected for 3 to 8 h after dosing from group 2 subjects. There were no serious adverse events or discontinuations due to adverse effects. In plasma, apixaban was the major circulating component and O-demethyl apixaban sulfate, a stable and water-soluble metabolite, was the significant metabolite. The exposure of apixaban (C(max) and area under the plasma concentration versus time curve) in subjects with bile collection was generally similar to that in subjects without bile collection. The administered dose was recovered in feces (group 1, 56.0%; group 2, 46.7%) and urine (group 1, 24.5%; group 2, 28.8%), with the parent drug representing approximately half of the recovered dose. Biliary excretion represented a minor elimination pathway (2.44% of the administered dose) from group 2 subjects within the limited collection period. Metabolic pathways identified for apixaban included O-demethylation, hydroxylation, and sulfation of hydroxylated O-demethyl apixaban. Thus, apixaban is an orally bioavailable inhibitor of factor Xa with elimination pathways that include metabolism and renal excretion.
Aim
Apixaban is an orally active inhibitor of coagulation factor Xa and is eliminated by multiple pathways, including renal and non‐renal elimination. Non‐renal elimination pathways consist of ...metabolism by cytochrome P450 (CYP) enzymes, primarily CYP3A4, as well as direct intestinal excretion. Two single sequence studies evaluated the effect of ketoconazole (a strong dual inhibitor of CYP3A4 and P‐glycoprotein P‐gp) and diltiazem (a moderate CYP3A4 inhibitor and a P‐gp inhibitor) on apixaban pharmacokinetics in healthy subjects.
Method
In the ketoconazole study, 18 subjects received apixaban 10 mg on days 1 and 7, and ketoconazole 400 mg once daily on days 4–9. In the diltiazem study, 18 subjects received apixaban 10 mg on days 1 and 11 and diltiazem 360 mg once daily on days 4–13.
Results
Apixaban maximum plasma concentration and area under the plasma concentration–time curve extrapolated to infinity increased by 62% (90% confidence interval CI, 47, 78%) and 99% (90% CI, 81, 118%), respectively, with co‐administration of ketoconazole, and by 31% (90% CI, 16, 49%) and 40% (90% CI, 23, 59%), respectively, with diltiazem.
Conclusion
A 2‐fold and 1.4‐fold increase in apixaban exposure was observed with co‐administration of ketoconazole and diltiazem, respectively.
The well accepted "free drug hypothesis" for small-molecule drugs assumes that only the free (unbound) drug concentration at the therapeutic target can elicit a pharmacologic effect. Unbound (free) ...drug concentrations in plasma are readily measurable and are often used as surrogates for the drug concentrations at the site of pharmacologic action in pharmacokinetic-pharmacodynamic analysis and clinical dose projection in drug discovery. Furthermore, for permeable compounds at pharmacokinetic steady state, the free drug concentration in tissue is likely a close approximation of that in plasma; however, several factors can create and maintain disequilibrium between the free drug concentration in plasma and tissue, leading to free drug concentration asymmetry. These factors include drug uptake and extrusion mechanisms involving the uptake and efflux drug transporters, intracellular biotransformation of prodrugs, membrane receptor-mediated uptake of antibody-drug conjugates, pH gradients, unique distribution properties (covalent binders, nanoparticles), and local drug delivery (e.g., inhalation). The impact of these factors on the free drug concentrations in tissues can be represented by
, the ratio of free drug concentration between tissue and plasma at steady state. This review focuses on situations in which free drug concentrations in tissues may differ from those in plasma (e.g.,
> or <1) and discusses the limitations of the surrogate approach of using plasma-free drug concentration to predict free drug concentrations in tissue. This is an important consideration for novel therapeutic modalities since systemic exposure as a driver of pharmacologic effects may provide limited value in guiding compound optimization, selection, and advancement. Ultimately, a deeper understanding of the relationship between free drug concentrations in plasma and tissues is needed.
The bioactivation of drugs is often associated with toxicological outcomes; however, for most cases, the causal relationship between bioactivation and toxicity is not well established despite ...extensive research that attempts to elucidate the mechanisms leading to the formation of chemically reactive species, presumably the initial step towards adverse reactions. Due to rapid advancement in the research of cytochrome P450s (CYPs) and the prevalence of CYP involvement in the metabolic clearance of pharmaceuticals, CYP-mediated bioactivation is widely investigated and reviewed, while non-CYP-mediated bioactivation has not been emphasized. The widespread use of metabolic stability screening in drug discovery, however, has led to the identification of new chemical entities that rely on non-CYP enzymes for clearance, and the number of drugs that undergo metabolism via these enzymes has increased. Non-CYP enzymes can be divided into four general categories according to their enzymatic function, namely, oxidative, reductive, conjugative and hydrolytic. The aim of this review is to complement the existing literature on CYP-mediated metabolism by focusing on bioactivation mediated non-CYP enzymes and provide representative examples in each category.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Background
Activated charcoal is commonly used to manage overdose or accidental ingestion of medicines. This study evaluated the effect of activated charcoal on apixaban exposure in human subjects.
...Methods
This was an open-label, three-treatment, three-period, randomized, crossover study of single-dose apixaban (20 mg) administered alone and with activated charcoal given at 2 or 6 h post-dose to healthy subjects. Blood samples for assay of plasma apixaban concentration were collected up to 72 h post-dose. Pharmacokinetic parameters, including peak plasma concentration (
C
max
), time to
C
max
(
T
max
), area under the concentration–time curve from time 0 to infinity (AUC
INF
), and terminal half-life (
T
½
), were derived from apixaban plasma concentration–time data. A general linear mixed-effect model analysis of
C
max
and AUC
INF
was performed to estimate the effect of activated charcoal on apixaban exposure.
Results
A total of 18 subjects were treated and completed the study. AUC
INF
for apixaban without activated charcoal decreased by 50 and 28 %, respectively, when charcoal was administered at 2 and 6 h post-dose. Apixaban
C
max
and
T
max
were similar across treatments. The mean
T
½
for apixaban alone (13.4 h) decreased to ~5 h when activated charcoal was administered at 2 or 6 h post-dose. Overall, apixaban was well tolerated in this healthy population, and most adverse events were consistent with the known profile of activated charcoal.
Conclusion
Administration of activated charcoal up to 6 h after apixaban reduced apixaban exposure and facilitated the elimination of apixaban. These results suggest that activated charcoal may be useful in the management of apixaban overdose or accidental ingestion.
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