Minor Elimination Pathway Research Articles

A phase 1 study to assess the absolute bioavailability, mass balance, pharmacokinetics, metabolism, and excretion of [14C]-mobocertinib, an oral inhibitor of EGFR exon 20 insertion mutations, in healthy participants

Mobocertinib (TAK-788) is a first-in-class oral epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor that received accelerated approval for the treatment of patients with non-small cell lung cancer with EGFR exon 20 insertion mutations previously treated with platinum-based chemotherapy. This phase 1, 2-period, study was conducted to assess the absolute bioavailability of mobocertinib (Period 1), as well as mass balance, pharmacokinetics, metabolism, and excretion of [14C]-mobocertinib (Period 2) in healthy adult males. In Period 1, participants received a single oral capsule dose of 160 mg mobocertinib, followed by a 15-minute intravenous infusion of 50 µg (~ 2 µCi) [14C]-mobocertinib administered from 3.75 to 4 h after the capsule dose. In Period 2, a single oral dose of 160 mg (~ 100 µCi) [14C]-mobocertinib was administered as an oral solution. The geometric mean absolute bioavailability of mobocertinib was determined to be 36.7%. After oral administration of [14C]-mobocertinib, mobocertinib and its active metabolites, AP32960 and AP32914, were minor components in plasma, accounting for only 0.275% of total plasma radioactivity as the majority of mobocertinib-related material was covalently bound to plasma proteins. The geometric mean percentage of the administered radioactive dose recovered in the urine and feces was 3.57% and 76.0%, respectively. Only 0.39% of the oral dose of [14C]-mobocertinib was recovered in the urine as mobocertinib; thus, indicating that renal excretion of unchanged drug was a very minor pathway of elimination. In both treatment periods, mobocertinib was generally safe and well-tolerated as all adverse events were Grade 1 in severity. (Trial registration number ClinicalTrials.gov NCT03811834. Registration date January 22, 2019).

Leveraging the mechanism of action (MOA) of xevinapant, an IAP inhibitor, for phase 3 dosing (RP3D) strategies in combination with radiation (RT) and chemoradiotherapy (CRT) in patients with locally advanced squamous cell carcinoma of the head and neck (LA SCCHN).

e18001 Background: Xevinapant is a first-in-class, oral IAP (inhibitor of apoptosis protein) inhibitor designed to restore cancer cell sensitivity to apoptosis, thereby enhancing the efficacy of RT and CRT. Xevinapant 200 mg/d (d1-14, Q3W) + CRT showed clinical benefit vs placebo + CRT in a phase (ph) 2 study of patients (pts) with LA SCCHN (NCT02022098). Exposure-response (E-R) relationships for efficacy and the composite safety endpoint of “mucositis and/or dysphagia” (AE_MD, a known RT-associated toxicity) were also identified. Two ph 3 studies of 200 mg/d xevinapant (d1-14, Q3W) + RT (XRay Vision; NCT05386550) or CRT (TrilynX; NCT04459715) followed by xevinapant monotherapy (MT) in pts with LA SCCHN are ongoing. We leverage the MOA of xevinapant to confirm RP3D strategy. Methods: Results from clinical and preclinical studies and quantitative pharmacology modeling were integrated. Results: Updated analyses of NCT02022098 data show an association between complete response (CR) and AE_MD in both treatment arms; AE_MD was the only AE with identified E-R relationship out of 13 AEs tested. In the xevinapant arm, the CR+AE_MD+ group had the highest median AUC but it overlapped with other groups (CR-AE_MD+, CR+AE_MD-, CR-AE_MD-) which all had similar AUCs. These data suggest that AE_MD is related to the RT-enhancing MOA of xevinapant and that decoupling efficacy from AE_MD incidence by exposure modulation is not feasible. Intermittent dosing for xevinapant was designed for recovery from epithelial toxicities considering the turnover of mucosal cells. In a human breast cancer mouse model, cIAP1 degradation with the intermittent regimen (100 mg/kg Q3D) at the end of the dosing interval was ~50% compared with the continuous one (30 mg/kg QD) expected to maintain maximal (max) cIAP1 degradation, but showed similar efficacy. This suggests continuous max cIAP1 degradation may not be required for efficacy, consistent with RT-enhancing MOA. Based on population PKPD modeling, at the RP3D for xevinapant max cIAP1 degradation was achieved during the dosing period; 91% of pts had at least 50% suppression at the end of the cycle. Xevinapant MT cycles are supported by enhanced anti-tumor activity with extended MT dosing following xevinapant/ RT cycles in MC38 syngeneic mouse model. Food-agnostic dosing is supported by similar AUCs observed in fed and fasted conditions. A human ADME study showed that urinary excretion is a minor elimination pathway, supporting no dose adjustment for pts with moderate renal impairment. Conclusions: Integrated assessment of all available clinical and nonclinical data supports the proposed RP3D strategy for xevinapant combined with RT or CRT, selected to maximize efficacy while minimizing potential adverse events and cumulative toxicity. Clinical trial information: NCT02022098 .

AZD5634, an inhaled ENaC inhibitor, in healthy subjects and patients with cystic fibrosis

BackgroundEpithelial sodium channel (ENaC) inhibitors may offer clinical benefit in cystic fibrosis (CF); however, data are limited. We report the outcomes of a Phase I (NCT02679729) and a Phase Ib (NCT02950805) study of AZD5634, a novel inhaled ENaC inhibitor. MethodsA Phase I, first-in-human, single-blind, placebo-controlled, single ascending dose, sequential dose group study assessed the safety, tolerability, and pharmacokinetics of AZD5634 in healthy subjects (n=53) in part A following inhaled doses up to 1700 µg, and, in part B, following administration of single inhaled (1700 µg) and intravenous (65 µg) doses. A Phase Ib, randomized, double-blind, placebo-controlled, single-dose, 2-way cross-over study assessed the effects of a single dose (600 µg) of inhaled AZD5634 on mucociliary clearance (MCC), pharmacokinetics and safety and tolerability in patients with CF (n=11). Nasal potential difference (NPD) was assessed as an in situ target engagement exploratory biomarker. ResultsAbsolute bioavailability of AZD5634 after inhalation was approximately 3%, indicating minimal distribution into the systemic circulation. Urinary excretion was a minor elimination pathway. Administration of inhaled AZD5634 did not improve MCC in CF patients, but AZD5634 inhibited ENaC in the nasal epithelium, as measured by NPD. AZD5634 was safe and well tolerated in both studies. ConclusionsAZD5634 showed favorable pharmacokinetics and safety in healthy subjects and patients with CF. However, despite achieving target engagement, proof of mechanism was not achieved after a single dose in patients with CF. Further evaluation into multiple dose studies is warranted to explore its therapeutic potential.

Characterization of the absorption, metabolism, excretion, and mass balance of gefapixant in humans.

Gefapixant (MK‐7264) is a first‐in‐class, selective antagonist of the P2X3 purinergic receptor currently being investigated as a therapeutic agent for the treatment of refractory or unexplained chronic cough. In non‐clinical studies, gefapixant was eliminated primarily by renal excretion of the parent drug. The objective of this study was to assess the disposition of gefapixant in humans. The absorption, metabolism, and excretion profiles of gefapixant were assessed after oral administration of a single dose of [14C]gefapixant to six healthy adult males. Following a single‐oral [14C]gefapixant dose to healthy adult males, the mass balance was achieved, with 98.9% of the administered radioactivity recovered in urine and feces. Elimination of gefapixant occurred primarily via renal excretion of the intact drug (64%); metabolism was a minor pathway of elimination of gefapixant (12% and 2% recovered in urine and feces, respectively). Single‐dose administration of [14C]gefapixant 50 mg was generally well tolerated in healthy adult males. The fraction of the anticipated therapeutic oral dose of gefapixant absorbed is estimated to be at least 78%. Gefapixant is expected to be the major circulating drug‐related material in plasma, and the majority of the dosed drug will be excreted unchanged in urine.

Combating antimicrobial resistance with cefiderocol for complicated infections involving the urinary tract.

Cefiderocol is a unique siderophore cephalosporin antimicrobial agent that has shown promise in treating complicated urinary tract infections (cUTI). Urinary tract infections are commonly diagnosed infections with risk increasing with age and prevalence more common in women. cUTI poses a risk of recurrence and is more likely to be associated with antibiotic-resistant bacteria. The Food and Drug Administration approved cefiderocol for use as a last-line option in the treatment of cUTI including pyelonephritis. Cefiderocol has activity against all forms of carbapenemases due to its ability to overcome the mechanisms of carbapenemase resistance. Because of this, resistance to cefiderocol is unlikely to occur. Studies show cefiderocol is well tolerated among younger patients and patients greater than 65 years of age, the latter making up most of the study population. Renal dose adjustments are recommended. Dose adjustment in the presence of hepatic impairment is not recommended, as hepatic clearance represents a minor elimination pathway for cefiderocol. The ability of cefiderocol to overcome multiple resistance mechanisms makes it a novel choice in combating multidrug-resistant bacteria in the treatment of cUTI.

DPP-4 Inhibitors: Renoprotective Potential and Pharmacokinetics in Type 2 Diabetes Mellitus Patients with Renal Impairment.

The continuously increasing incidence of diabetes worldwide has attracted the attention of the scientific community and driven the development of a novel class of antidiabetic drugs that can be safely and effectively used in diabetic patients. Of particular interest in this context are complications associated with diabetes, such as renal impairment, which is the main cause of high cardiovascular morbidity and mortality in diabetic patients. Intensive control of glucose levels and other risk factors associated with diabetes and metabolic syndrome provides the foundations for both preventing and treating diabetic nephropathy. Dipeptidyl peptidase-4 (DPP-4) inhibitors represent a highly promising novel class of oral agents used in the treatment of type 2 diabetes mellitus that may be successfully combined with currently available antidiabetic therapeutics in order to achieve blood glucose goals. Beyond glycemic control, emerging evidence suggests that DPP-4 inhibitors may have desirable off-target effects, including renoprotection. All type 2 diabetes mellitus patients with impaired renal function require dose adjustment of any DPP-4 inhibitor administered except for linagliptin, for which renal excretion is a minor elimination pathway. Thus, linagliptin is the drug most frequently chosen to treat type 2 diabetes mellitus patients with renal failure.

Preclinical Pharmacokinetics Study of R- and S-Enantiomers of the Histone Deacetylase Inhibitor, AR-42 (NSC 731438), in Rodents.

AR-42, a new orally bioavailable, potent, hydroxamate-tethered phenylbutyrate class I/IIB histone deacetylase inhibitor currently is under evaluation in phase 1 and 2 clinical trials and has demonstrated activity in both hematologic and solid tumor malignancies. This report focuses on the preclinical characterization of the pharmacokinetics of AR-42 in mice and rats. A high-performance liquid chromatography-tandem mass spectrometry assay has been developed and applied to the pharmacokinetic study of the more active stereoisomer, S-AR-42, when administered via intravenous and oral routes in rodents, including plasma, bone marrow, and spleen pharmacokinetics (PK) in CD2F1 mice and plasma PK in F344 rats. Oral bioavailability was estimated to be 26 and 100% in mice and rats, respectively. R-AR-42 was also evaluated intravenously in rats and was shown to display different pharmacokinetics with a much shorter terminal half-life compared to that of S-AR-42. Renal clearance was a minor elimination pathway for parental S-AR-42. Oral administration of S-AR-42 to tumor-bearing mice demonstrated high uptake and exposure of the parent drug in the lymphoid tissues, spleen, and bone marrow. This is the first report of the pharmacokinetics of this novel agent, which is now in early phase clinical trials.

Comment on: “Enzalutamide: A Step Towards Pharmacokinetic-Based Dosing in Men with Metastatic Castration-Resistant Prostate Cancer”

Dear Editors, We thank Dr. Paul Hutson for his thorough commentary [1] on our recent article entitled ‘Enzalutamide: a step towards pharmacokinetic-based dosing in men with metastatic castration-resistant prostate cancer’ [2]. We would like to provide an important correction to Dr. Hutson’s commentary, regarding the following statements: Gibbons et al. do not report the renal clearance or the effect of renal failure on the clearance of enzalutamide or its active N-desmethyl metabolite, and to date no other reports of enzalutamide renal clearance are available to guide dosing. The mass-balance studies in the present study demonstrate that approximately 60 % of the dose was recovered in the urine, suggesting that dosing of the drug may need to be reduced in patients with kidney dysfunction. While not emphasized, the article does describe renal clearance on page 1050 (paragraph 2): The major route of excretion of 14C-radioactivity was urine (71.0 % of dose), primarily as metabolites. The remainder of the radioactivity was excreted in feces (13.6 % of dose). Based on LC–MS/MS analysis, the main component in urine was the carboxylic acid metabolite, which accounted for 62.7 % of the dose. A trace amount of unchanged parent enzalutamide was excreted in urine. N-Desmethyl enzalutamide was too low to quantitate in urine by LC–MS/MS. Based on this information, renal excretion is a minor elimination pathway for unchanged parent enzalutamide and N-desmethyl enzalutamide. As indicated in this excerpt, 62.7 % of the dose was recovered in urine as the inactive carboxylic acid metabolite, and the remainder was additional minor metabolites; enzalutamide and N-desmethyl enzalutamide (active metabolite) were present in urine at only trace amounts. These data indicate that renal excretion is a minor elimination pathway for unchanged parent enzalutamide and N-desmethyl enzalutamide. Further details are captured in the US prescribing information for XTANDI® (enzalutamide) capsules (product label), which states the following: A dedicated renal impairment trial for XTANDI has not been conducted. Based on the population pharmacokinetic analysis using data from clinical trials in patients with metastatic CRPC and healthy volunteers, no significant difference in enzalutamide clearance was observed in patients with pre-existing mild to moderate renal impairment [30 mL/min ≤ creatinine clearance (CrCL) ≤ 89 mL/min] compared to patients and volunteers with baseline normal renal function (CrCL ≥90 mL/min). No initial dosage adjustment is necessary for patients with mild to moderate renal impairment. Severe renal impairment (CrCL <30 mL/min) and end-stage renal disease have not been assessed [see Clinical Pharmacology (12.3)]. In summary, data presented in our original research article [2], as well as results in the XTANDI® product label, indicate that starting dose reductions are not required in patients with mild to moderate renal impairment. There are no data to assess the effects of severe renal impairment (CrCL <30 mL/min). As Dr. Hutson’s commentary misstates dosing recommendations for XTANDI®, we request that you kindly publish a supplementary response to Dr. Hutson’s commentary in Clinical Pharmacokinetics.

Deletion ofAbcg2Has Differential Effects on Excretion and Pharmacokinetics of Probe Substrates in Rats

This study was designed to characterize breast cancer resistance protein (Bcrp) knockout Abcg2(-/-) rats and assess the effect of ATP-binding cassette subfamily G member 2 (Abcg2) deletion on the excretion and pharmacokinetic properties of probe substrates. Deletion of the target gene in the Abcg2(-/-) rats was confirmed, whereas gene expression was unaffected for most of the other transporters and metabolizing enzymes. Biliary excretion of nitrofurantoin, sulfasalazine, and compound A [2-(5-methoxy-2-((2-methyl-1,3-benzothiazol-6-yl)amino)-4-pyridinyl)-1,5,6,7-tetrahydro-4H-pyrrolo[3,2-c]pyridin-4-one] accounted for 1.5, 48, and 48% of the dose in the Abcg2(+/+) rats, respectively, whereas it was decreased by 70 to 90% in the Abcg2(-/-) rats. Urinary excretion of nitrofurantoin, a significant elimination pathway, was unaffected in the Abcg2(-/-) rats, whereas renal clearance of sulfasalazine, a minor elimination pathway, was reduced by >90%. Urinary excretion of compound A was minimal. Systemic clearance in the Abcg2(-/-) rats decreased 22, 43 (p<0.05), and 57%, respectively, for nitrofurantoin, sulfasalazine, and compound A administered at 1 mg/kg and 27% for compound A administered at 5 mg/kg. Oral absorption of nitrofurantoin, a compound with high aqueous solubility and good permeability, was not limited by Bcrp. In contrast, the absence of Bcrp led to a 33- and 11-fold increase in oral exposure of sulfasalazine and compound A, respectively. These data show that Bcrp plays a crucial role in biliary excretion of these probe substrates and has differential effects on systemic clearance and oral absorption in rats depending on clearance mechanisms and compound properties. The Abcg2(-/-) rat is a useful model for understanding the role of Bcrp in elimination and oral absorption.

Apixaban Metabolism and Pharmacokinetics after Oral Administration to Humans

The metabolism and disposition of [(14)C]apixaban, 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.

Population Pharmacokinetic/Pharmacodynamic Modelling of the Analgesic Effects of Tramadol in Pediatrics

The efficacy of tramadol (T) in children is not clearly understood because it is still unknown the ability of that population to form the active metabolite O-demethyltramadol (M1) and, whether or not the parent compound has a contribution to the efficacy. The aim was to develop a population pharmacokinetic/pharmacodynamic model for T in pediatrics, identifying the main active components. One hundred four children, mean age (4.55 years) received intravenously 1 mg/kg dose of T over 2.5 min at the end of surgery. If pain relief was inadequate, then an additional 0.33 mg/kg dose was given at 15, 30 and/or 45 min. Plasma samples and analgesic responses such as crying and movement were measured during a 6-h period. The estimates of the apparent volumes of distribution of the central compartment and at steady state and total plasma clearance of T were 8 l, 46.2 l, and 15.2 l/h, respectively. M1 formation clearance represented only a minor elimination pathway of T. Effect site concentrations of T and M1 were found to be the best predictors of the movement and crying responses, respectively. Steady-state plasma concentration levels of T and M1 of 100 and 15 ng/ml were associated with a 95% probability of adequate pain relief. Children have the ability to produce enough M1 to achieve proper pain relief. The response variables investigated give further evidence that not only the opioid effects of the metabolite are relevant, also the non-opiod effects of tramadol seem to give a significant contribution in its clinical use.

Disposition of [14C]Ruboxistaurin in Humans

Ruboxistaurin is a potent and specific inhibitor of the beta isoforms of protein kinase C (PKC) that is being developed for the treatment of diabetic microvascular complications. The disposition of [(14)C]ruboxistaurin was determined in six healthy male subjects who received a single oral dose of 64 mg of [(14)C]ruboxistaurin in solution. There were no clinically significant adverse events during the study. Whole blood, urine, and feces were collected at frequent intervals after dosing. Metabolites were profiled by high performance liquid chromatography with radiometric detection. The total mean recovery of the radioactive dose was approximately 87%, with the majority of the radioactivity (82.6 +/- 1.1%) recovered in the feces. Urine was a minor pathway of elimination (4.1 +/- 0.3%). The major route of ruboxistaurin metabolism was to the N-desmethyl ruboxistaurin metabolite (LY338522), which has been shown to be active and equipotent to ruboxistaurin in the inhibition of PKC(beta). In addition, multiple hydroxylated metabolites were identified by liquid chromatography-mass spectrometry in all matrices. Pharmacokinetics were conducted for both ruboxistaurin and LY338522 (N-desmethyl ruboxistaurin, 1). These moieties together accounted for approximately 52% of the radiocarbon measured in the plasma. The excreted radioactivity was profiled using radiochromatography, and approximately 31% was structurally characterized as ruboxistaurin or N-desmethyl ruboxistaurin. These data demonstrate that ruboxistaurin is metabolized primarily to N-desmethyl ruboxistaurin (1) and multiple other oxidation products, and is excreted primarily in the feces.

Toxicokinetics and biotransformation of 3-(4-methylbenzylidene)camphor in rats after oral administration

3-(4-Methylbenzylidene)camphor (4-MBC) is an UV-filter frequently used in sunscreens and cosmetics. Equivocal findings in some screening tests for hormonal activity initiated a discussion on a possible weak estrogenicity of 4-MBC. In this study, the toxicokinetics and biotransformation of 4-MBC were characterized in rats after oral administration. Male and female Sprague-Dawley rats (n = 3 per group) were administered single oral doses of 25 or 250 mg/kg bw of 4-MBC in corn oil. Metabolites formed were characterized and the kinetics of elimination for 4-MBC and its metabolites from blood and with urine were determined. Metabolites of 4-MBC were characterized by (1)H NMR and LC-MS/MS as 3-(4-carboxybenzylidene)camphor and as four isomers of 3-(4-carboxybenzylidene)hydroxycamphor containing the hydroxyl group located in the camphor ring system with 3-(4-carboxybenzylidene)-6-hydroxycamphor as the major metabolite. After oral administration of 4-MBC, only very low concentrations of 4-MBC were present in blood and the peak concentrations of 3-(4-carboxybenzylidene)camphor were approximately 500-fold above those of 4-MBC; blood concentrations of 3-(4-carboxybenzylidene)-6-hydroxycamphor were below the limit of detection. Blood concentration of 4-MBC and 3-(4-carboxybenzylidene)camphor peaked within 10 h after 4-MBC administration and then decreased with half-lives of approximately 15 h. No major differences in peak blood levels between male and female rats were seen. In urine, one isomer of 3-(4-carboxybenzylidene)hydroxycamphor was the predominant metabolite [3-(4-carboxybenzylidene)-6-hydroxycamphor], the other isomers and 3-(4-carboxybenzylidene)camphor were only minor metabolites excreted with urine. However, urinary excretion of 4-MBC-metabolites represents only a minor pathway of elimination for 4-MBC, since most of the applied dose was recovered in feces as 3-(4-carboxybenzylidene)camphor and, to a smaller extent, as 3-(4-carboxybenzylidene)-6-hydroxycamphor. Glucuronides of both metabolites were also present in feces, but partly decomposed during sample workup and were thus not quantified. The results show that absorbed 4-MBC undergoes extensive first-pass biotransformation in rat liver resulting in very low blood levels of the parent 4-MBC. Enterohepatic circulation of glucuronides derived from the two major 4-MBC metabolites may explain the slow excretion of 4-MBC metabolites with urine and the small percentage of the administered doses recovered in urine.

Factors affecting the pharmacokinetic profile of MS-275, a novel histone deacetylase inhibitor, in patients with cancer

To evaluate elimination pathways of the histone deacetylase inhibitor MS-275 in vitro and screen for relationships between demographic factors that may affect its pharmacokinetics in vivo. Substrate specificity of MS-275 for the liver-specific organic anion transporting polypeptides (OATPs) was assessed using Xenopus laevis oocytes, and in vitro metabolism was evaluated using human liver microsomes. In vivo pharmacokinetic data were obtained from 64 adult patients (36 male/28 female; median age, 57 years) receiving MS-275 orally (dose range, 2 to 12 mg/m2). Accumulation of [G-3H]MS-275 by oocytes expressing OATP1B1 or OATP1B3 was not significantly different from water-injected controls (p = 0.82). Furthermore, no metabolites could be detected after incubation of MS-275 in human liver microsomes, suggesting that hepatic metabolism is a minor pathway of elimination. The mean (+/- SD) apparent oral clearance of MS-275 was 38.5 +/- 18.7 L/h, with a coefficient of variation (%CV) of 48.7%. When clearance was adjusted for body-surface area (BSA), the inter-individual variability was similar (%CV = 50.1%). In addition, in a linear-regression analysis, except for adjusted ideal body weight (p = 0.02, |r| = 0.29), none of the studied measures (BSA, lean-body mass, ideal body weight, body-mass index, height, weight, age, and sex) was a significant covariate (p > 0.13; |r| < 0.11) for oral clearance. The current analysis has eliminated a number of candidate covariates from further consideration as important determinants of MS-275 absorption and disposition. Furthermore, MS-275 can be added to the list of cancer drugs where BSA-based dosing is not more accurate than fixed dosing.

Comparison of pharmacokinetics and tissue disposition of an antisense phosphorothioate oligonucleotide targeting human Ha‐ras mRNA in mouse and monkey

The plasma pharmacokinetics and tissue disposition of ISIS 2503 were studied in mice following single and multiple bolus intravenous (iv) injections of 1–50 mg/kg, and in monkeys following single and multiple 2‐h iv infusions of 1–10 mg/kg and bolus iv injections of 1 mg/kg of ISIS 2503. ISIS 2503 and its metabolites were measured in plasma, urine, and tissues using solid‐phase extraction followed by capillary gel electrophoresis (CGE). In both species, the plasma clearance of ISIS 2503 was characterized by rapid distribution to tissues, and to a lesser extent, metabolism. The plasma clearance in mice was at least two‐fold more rapid than in monkeys at equivalent doses. The plasma disposition (t1/2) increased with dose. The highest concentrations of oligonucleotide were consistently observed in the kidney and liver in both species. At equivalent doses, tissue concentrations in monkeys were much higher than tissue concentrations in mice. Urinary excretion of total oligonucleotide was a minor elimination pathway in both species at doses < 10 mg/kg. However, urinary excretion of total oligonucleotide in mice was increased to 12–29% as dose increased from 20 to 50 mg/kg. © 2001 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 90:182–193, 2001

In vivo stability, disposition and metabolism of a “hybrid” oligonucleotide phosphorothioate in rats

Oligodeoxynucleotide phosphorothioates containing segments of 2′- O-methyloligoribo-nucleotide phosphorothioates at both 3′- and 5′-ends (hybrid oligonucleotide) have been shown to be potent antisense agents. In the present study, in vivo biostability, disposition, and excretion of a 25-mer hybrid oligonucleotide were determined in rats after i.v. bolus administration of the 35S-labeled oligonucleotide at a dose of 30 mg/kg. The plasma disappearance curve for the hybrid oligonucleotide could be described by a two-compartmental model, with half-lives of 0.34 and 52.02 hr, respectively. The majority of the radioactivity in plasma was associated with the intact hybrid oligonucleotide. Urinary excretion represented the major pathway of elimination, with 21.98 ± 3.21% (mean ± SD) of the administered dose excreted within 24 hr and 38.13 ± 2.99% over 240 hr post-dosing. The majority of the radioactivity in urine was associated with the degradative products with lower molecular weights, but the intact form was also detected by HPLC analysis. Fecal excretion was a minor pathway of elimination with 2.34 ± 0.13% of the administered dose excreted over 24 hr and 6.74 ± 0.40% over 240 hr post-dosing. A wide tissue distribution of hybrid oligonucleotide was observed based on radioactivity levels, and analysis by HPLC showed that the majority of the radioactivity in tissues was associated with the intact hybrid oligonucleotide. Further analyses of the experimental data provided a comprehensive pharmacokinetic analysis of hybrid oligonucleotide in each tissue. Compared with a previously examined oligodeoxynucleotide phosphorothioate (GEM 91) that has a similar nucleotide sequence, the hybrid oligonucleotide had a shorter distribution half-life and a longer elimination half-life, based on the quantitation of radioactivity in plasma. Although it had a similar tissue distribution pattern compared with other oligonucleotide phosphorothioates such as GEM 91, the hybrid oligonucleotide was more stable in vivo, which may be important in the development of antisense oligonucleotides as therapeutic agents.

In vivo stability and disposition of a self-stabilized oligodeoxynucleotide phosphorothioate in rats

The use of antisense oligonucleotides represents a novel, genetically based therapy. The biostability and pharmacokinetics of a 33-mer self-stabilized oligodeoxynucleotide with significant anti-HIV activity was determined in rats after intravenous administration of [35S]oligodeoxynucleotide. Plasma disappearance of the labeled oligodeoxynucleotide could be described by a two-compartment model, with half-lives of 0.54 and 41.44 h. The oligodeoxynucleotide in plasma remained mainly intact. Urinary excretion represented the major elimination pathway, with approximately 27% of the administered dose excreted within 24 h and 57% over 240 h. The majority of radioactivity in urine was attached to degradative products. Fecal excretion was a minor elimination pathway. A wide tissue distribution of the oligonucleotide was observed, with the majority of radioactivity in most tissues being intact. Compared with other linear oligonucleotide phosphorothioates, the self-stabilized oligonucleotide was more stable in vivo, which may be important in development of antisense oligonucleotides as therapeutic agents.

Pharmacokinetics and tissue distribution in rats of an oligodeoxynucleotide phosphorothioate (GEM 91) developed as a therapeutic agent for human immunodeficiency virus type-1

An antisense oligodeoxynucleotide phosphorothioate, namely gene expression modulator 91 (GEM 91), has been demonstrated to have significant anti-human immunodeficiency virus activity in various tissue culture models. The present study was undertaken to determine the pharmacokinetics and tissue distribution of GEM 91 in rats following i.v. bolus administration of 35S-radiolabeled GEM 91. Plasma disappearance curves for GEM 91-derived radioactivity could be described by the sum of two exponentials, with half-lives (mean ± SEM) of 0.95 (± 0.07) and 47.57 (± 14.48) hr. Urinary excretion represented the major pathway of elimination of GEM 91, with 26.67 ± 6.46% (mean ± SD) of the administered dose excreted within 24 hr and 58.12 ± 4.36% over 240 hr after GEM 91 administration. Fecal excretion was a minor pathway of elimination of GEM 91 with 1.4 ± 0.62% (mean ± SD) of the administered dose excreted over 24 hr and 8.54 ± 0.64% over 240 hr. A wide tissue distribution of GEM 91 was observed. During the initial 30 min, the highest levels of tissue radioactivity were found in the kidney, liver, spleen, lungs, and heart. Radioactivity was retained over longer time periods in the kidneys, liver, heart, and intestine. Analyses of the extracted radioactivities from plasma, kidney, and liver by gel electrophoresis showed the presence of both intact GEM 91 and degradative products with smaller molecular weights. Radioactivity in urine was found to be degradative metabolites of GEM 91. Based on the experimental data, pharmacokinetic parameters for GEM 91 in each tissue and biological fluids were calculated using computer-based two-compartmental i.v. bolus or absorption models. This study is important not only in providing the basis for future studies of GEM 91 in humans, but also in understanding the pharmacology and toxicology of antisense oligodeoxynucleotide phosphorothioates, in general.