10-fold Dose Range Research Articles

Abstract 49: LYL119, an investigational ROR1-targeted CAR T-cell product incorporating four novel reprogramming technologies designed for effective cell therapy for solid tumors

Abstract Effective solid tumor cell therapy requires new strategies to improve T-cell activation, persistence, and durable function. We developed four complementary T-cell reprogramming technologies to enhance chimeric antigen receptor (CAR) T-cell therapy in solid tumors: 1) overexpression of the activator protein 1 (AP-1) family transcription factor c-Jun to delay T-cell exhaustion and improve antitumor activity; 2) nuclear receptor subfamily 4A member 3 (NR4A3) gene knockout (KO) to further delay exhaustion and enhance functionality; 3) Epi-RTM manufacturing protocols to preserve stem-like characteristics; and 4) Stim-RTM technology, a novel activating reagent to improve product potency compared with standard reagents. LYL119 is an investigational ROR1-targeted CAR T-cell product that combines these technologies to create potent and durable CAR T cells. Healthy or NSCLC patient donor T cells were manufactured with the Epi-R protocols, activated with Stim-R or a standard reagent, and transduced with a vector encoding a ROR1 CAR and c-Jun. The NR4A3 gene or a control gene was edited using SpyFiTM Cas9 nuclease (Aldevron®). Cytotoxicity, cytokine production, phenotype, and single-cell transcriptomic and epigenetic profiles were evaluated in vitro after antigen restimulation assays designed to promote exhaustion. CAR T cell activity was evaluated in vivo using a ROR1+ NSCLC xenograft mouse model. Research and clinical scale LYL119 products achieved ~90% genome editing efficiency at the NR4A3 target gene resulting in a 13-fold protein reduction compared to non-edited CAR T cells. LYL119 exhibited superior cytotoxicity and cytokine production upon antigen restimulation across 7 different ROR1+ solid tumor cell lines compared to CAR T cells that lacked one or more reprogramming technologies. After repeated rounds of tumor cell killing, LYL119 displayed reduced surface expression of exhaustion-related receptors (e.g. TIM-3) and higher expression of stemness-related markers (e.g. CD127) compared to non-edited CAR T cells. Furthermore, transcriptomic analysis revealed global downregulation of exhaustion-related gene signatures and retention of unique cell subsets characterized by upregulation of memory and effector-associated gene signatures. LYL119 exhibited robust antitumor efficacy in vivo across a 10-fold dose range, including a very low dose of 1 × 105 CAR T cells. Lastly, LYL119 derived from NSCLC patient donor T cells also demonstrated enhanced cytotoxicity in vitro compared to control CAR T cells. These nonclinical data suggest LYL119, which combines c-Jun overexpression, NR4A3 KO, Epi-R protocols, and Stim-R technology, can limit exhaustion, maintain stem-like features, and has the potential to provide effective and durable CAR T-cell antitumor activity in patients with ROR1+ solid tumors. Citation Format: Viola C. Lam, Aileen Li, Meritxell Galindo Casas, Jessica Barragan, Christina Cheung, Jessica Briones, Esha Afreen, Grant Vavra, Jia Lu, Purnima Sundar, Rowena Martinez, Candace Sims, Shobha Potluri, Omar Ali, Alexander S. Cheung, Rachel C. Lynn. LYL119, an investigational ROR1-targeted CAR T-cell product incorporating four novel reprogramming technologies designed for effective cell therapy for solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 49.

PK/PD Modeling to Inform Clinical Development of an Adeno-associated Virus Gene Transfer Therapy for Duchenne Muscular Dystrophy (S48.005)

<h3>Objective:</h3> To evaluate the pharmacokinetic (PK)/pharmacodynamic (PD) relationship between tissue vector genome exposure, biological efficacy and functional outcome in an animal model of Duchenne muscular dystrophy (DMD) (DMD^mdx mice) following treatment with delandistrogene moxeparvovec (SRP-9001). <h3>Background:</h3> Delandistrogene moxeparvovec is an investigational gene transfer therapy developed to address the root cause of DMD through targeted skeletal and cardiac muscle expression of SRP-9001 dystrophin protein, which contains key functional domains of dystrophin. <h3>Design/Methods:</h3> We evaluated vector biodistribution, expression, and clinical dose selection of delandistrogene moxeparvovec using a novel application of a PK and PD modeling approach applied to data collected from DMD^mdx mice. We analyzed PK/PD relationships between dose, tissue vector genome exposure, SRP-9001 dystrophin protein expression (percent dystrophin positive fibers [PDPF] and western blot), and functional improvement (relative specific force from tibialis anterior and diaphragm). <h3>Results:</h3> Linear kinetics with a dose-proportional increase in tissue drug exposure were demonstrated across the nearly 10-fold dose range (4.43×10¹³–4.01×10¹⁴ vg/kg), and in all tissues. The relationship between tissue vector exposure and PD endpoints (PDPF, motor function outcome) showed a saturable response across a wide range of vector exposures. The vector exposure at 1.33×10¹⁴ vg/kg (the clinical dose) approached the maximum treatment response. Relative specific force and PDPF were significantly correlated (<i>P</i>=4.43×10⁻⁶). However, the relationship appeared to be nonlinear, with increased PDPF expression approaching the maximal functional improvement. Relative specific force and western blot were not significantly correlated. <h3>Conclusions:</h3> For the first time, biodistribution, biomarker and functional efficacy data were used to quantify and demonstrate PK/PD relationships for an adeno-associated virus (AAV)-based gene transfer therapy in a DMD animal model. The results continue to support the expected therapeutic benefit and clinical dose of delandistrogene moxeparvovec, an AAV-based gene transfer therapy. <b>Disclosure:</b> Mrs. East has received personal compensation for serving as an employee of Sarepta Therapeutics. Rachael Potter has received stock or an ownership interest from Sarepta Therapeutics. Dr. Snedeker has received personal compensation for serving as an employee of Sarepta Therapeutics. Dr. Snedeker has stock in Sarepta Therapeutics. Dr. Haile has nothing to disclose. Christopher Wier has received personal compensation for serving as an employee of Sarepta Therapeutics. Christopher Wier has stock in Sarepta Therapeutics. Ms. Rodino-Klapac has received personal compensation for serving as an employee of Sarepta Therapeutics, Inc.. Ms. Rodino-Klapac has received stock or an ownership interest from Sarepta Therapeutics. Ms. Rodino-Klapac has received intellectual property interests from a discovery or technology relating to health care.

VP.82 PK/PD modelling to inform clinical development of an adeno-associated virus gene transfer therapy for Duchenne muscular dystrophy

Delandistrogene moxeparvovec (SRP-9001) is an investigational gene transfer therapy for Duchenne muscular dystrophy (DMD) developed for targeted skeletal and cardiac muscle expression of micro-dystrophin—a shortened, functional dystrophin protein. We evaluated vector biodistribution, expression, and clinical dose selection of delandistrogene moxeparvovec using a novel application of a pharmacokinetic (PK) and pharmacodynamic (PD) modelling approach applied to data collected from an animal model of DMD (DMD^MDX mice). We analysed PK/PD relationships between dose, tissue vector genome exposure, micro-dystrophin protein expression (percentage dystrophin positive fibres [PDPF] and western blot), and functional improvement (relative specific force from tibialis anterior and diaphragm). Linear kinetics with a dose-proportional increase in tissue drug exposure was demonstrated across the nearly 10-fold dose range (4.43x10¹³–4.01x10¹⁴ vg/kg), and in all tissues. The relationship between tissue vector exposure and PD endpoints (PDPF, motor function outcome) showed a saturable response across a wide range of vector exposures. The vector exposure at 1.33x10¹⁴ vg/kg (the clinical dose) approached the maximum treatment response. Relative specific force and PDPF were significantly correlated (P=4.43x10^-6). However, the relationship appeared to be nonlinear, with increased PDPF expression approaching the maximal functional improvement. Relative specific force and western blot were not significantly correlated. For the first time, biodistribution, biomarker, and functional efficacy data were used to quantify and demonstrate PK/PD relationships for an adeno-associated virus (AAV)-based gene transfer therapy in a DMD animal model. The results continue to support the expected therapeutic benefit and clinical dose of delandistrogene moxeparvovec, an AAV-based gene transfer therapy.

Noncompartmental pharmacokinetics of three intravenous mycophenolate mofetil concentrations in healthy Standardbred mares.

Mycophenolate mofetil (MMF) is the prodrug of mycophenolic acid (MPA) which acts as an immunosuppressive agent. During the biotransformation of MMF to MPA, additional metabolites including MPA phenol glucuronide (MPAG), MPA acyl glucuronide (AcMPAG) and MPA phenol glucoside (MPG) are formed. To define the noncompartmental pharmacokinetic (PK) parameters of three single doses of intravenous (i.v.) MMF and its downstream metabolites in healthy horses. Six healthy Standardbred mares. Generic MMF (Par Pharmaceuticals; Chestnut Ridge, NY, USA) was reconstituted and administered as a single i.v. bolus at 1.0 mg/kg, 5.0 mg/kg and 10.0 mg/kg with an eight day washout between treatments. Blood samples were collected immediately before MMF administration and over 24 h. A liquid chromatography-tandem mass spectrometry assay was developed following FDA guidance to determine plasma MMF, MPA, MPAG, AcMPAG and MPG concentrations. Plasma concentrations were analysed independently, followed by calculation of geometric mean and coefficient of variation. Noncompartmental PK parameters were determined for MMF and all metabolites at all doses. MMF was rapidly converted to MPA in all horses. Each incremental dose of MMF resulted in increases in Cmax and AUCinf _obs for MPA and the three additional metabolites. Within the 10-fold dose range, the increase in Cmax and AUCinf _obs for MMF and its metabolites was nonlinear. Horses biotransform MMF into MPA, MPAG, AcMPAG and MPG via the glucuronidation and glucosidation clearance pathways. Equine reference PK profiles for MPA and the metabolites, MPAG, AcMPAG and MPG were established.

VIP152 Is a Novel CDK9 Inhibitor with Efficacy in Chronic Lymphocytic Leukemia

VIP152 Is a Novel CDK9 Inhibitor with Efficacy in Chronic Lymphocytic Leukemia

Response to mepolizumab treatment is sustained across 4-weekly dosing periods.

BackgroundMepolizumab (100 mg delivered s.c. every 4 weeks) is indicated for add-on maintenance treatment for patients with severe eosinophilic asthma. Mepolizumab has been shown to reduce exacerbations and the requirement for daily oral corticosteroids, and improve asthma control and symptoms. However, data on the durability of the response to mepolizumab during dosing periods are limited. The aim of this study was to investigate the efficacy profile in patients with severe eosinophilic asthma over the 4-weekly dosing period for various fixed mepolizumab doses.MethodsThis was a post hoc analysis of data from the phase IIb/III DREAM study. Patients ≥12 years of age with severe eosinophilic asthma were randomised (1:1:1:1) to receive intravenous mepolizumab 75 mg (equivalent to 100 mg s.c.), 250 mg, 750 mg or placebo, plus standard of care, every 4 weeks for 52 weeks. The number of exacerbations and eDiary data (peak expiratory flow, rescue medication use and symptom scores) from two periods in each 4-weekly dosing interval (days 1–14 and 15–28) over the 52-week treatment period were analysed.FindingseDiary data and the proportion of patients experiencing ≥1 exacerbation were similar during the first and second 2 weeks of a dosing period across all mepolizumab doses.InterpretationThese results demonstrate that the response to mepolizumab is sustained over the 4-weekly dosing period with no differences across a 10-fold dose range and supports the use of the current mepolizumab dosing regimen in patients with severe eosinophilic asthma.

Discovery of liver-directed glucokinase activator having anti-hyperglycemic effect without hypoglycemia

Glucokinase activators (GKAs) are among the emerging drug candidates for the treatment of type 2 diabetes (T2D). Despite effective blood glucose lowering in clinical trials, many pan-GKAs “acting both in pancreas and liver” have been discontinued from clinical development mainly because of their potential to cause hypoglycemia. Pan-GKAs over sensitize pancreatic GK, resulting in insulin secretion even at sub-normoglycemic level which might be a possible explanation for hypoglycemia. An alternative approach to minimize the risk of hypoglycemia is to use liver-directed GKAs, which are reported to be advancing well in clinical development. Here, we report the discovery and structure-activity relationship (SAR) studies on a novel 2-phenoxy-acetamide series with the aim of identifying a liver-directed GKA. Incorporation of a carboxylic acid moiety as an active hepatocyte uptake recognizing element at appropriate position of 2-phenoxy-acetamide core led to the identification of 26, a potent GKA with predominant liver-directed pharmacokinetics in mice. Compound 26 on oral administration significantly reduced blood glucose levels during an oral glucose tolerance test (oGTT) performed in diet-induced obese (DIO) mice, while showing no sign of hypoglycemia in normal C57 mice over a 10-fold dose range, even when dosed at fasted condition. Together, these data demonstrate a liver-directed GKA has beneficial effect on glucose homeostasis with reduced risk of hypoglycemia.

Use of microdosing and accelerator mass spectrometry to evaluate the pharmacokinetic linearity of a novel tricyclic GyrB/ParE inhibitor in rats.

Determining the pharmacokinetics (PKs) of drug candidates is essential for understanding their biological fate. The ability to obtain human PK information early in the drug development process can help determine if future development is warranted. Microdosing was developed to assess human PKs, at ultra-low doses, early in the drug development process. Microdosing has also been used in animals to confirm PK linearity across subpharmacological and pharmacological dose ranges. The current study assessed the PKs of a novel antimicrobial preclinical drug candidate (GP-4) in rats as a step toward human microdosing studies. Dose proportionality was determined at 3 proposed therapeutic doses (3, 10, and 30 mg/kg of body weight), and PK linearity between a microdose and a pharmacological dose was assessed in Sprague-Dawley rats. Plasma PKs over the 3 pharmacological doses were proportional. Over the 10-fold dose range, the maximum concentration in plasma and area under the curve (AUC) increased 9.5- and 15.8-fold, respectively. PKs from rats dosed with a (14)C-labeled microdose versus a (14)C-labeled pharmacological dose displayed dose linearity. In the animals receiving a microdose and the therapeutically dosed animals, the AUCs from time zero to infinity were 2.6 ng · h/ml and 1,336 ng · h/ml, respectively, and the terminal half-lives were 5.6 h and 1.4 h, respectively. When the AUC values were normalized to a dose of 1.0 mg/kg, the AUC values were 277.5 ng · h/ml for the microdose and 418.2 ng · h/ml for the pharmacological dose. This 1.5-fold difference in AUC following a 300-fold difference in dose is considered linear across the dose range. On the basis of the results, the PKs from the microdosed animals were considered to be predictive of the PKs from the therapeutically dosed animals.

Safety, pharmacokinetics and pharmacodynamics of multiple oral doses of apixaban, a factor Xa inhibitor, in healthy subjects

Apixaban is an oral factor Xa inhibitor approved for stroke prevention in atrial fibrillation and thromboprophylaxis in patients who have undergone elective hip or knee replacement surgery and under development for treatment of venous thromboembolism. This study examined the safety, pharmacokinetics and pharmacodynamics of multiple dose apixaban. This double-blind, randomized, placebo-controlled, parallel group, multiple dose escalation study was conducted in six sequential dose panels - apixaban 2.5, 5, 10 and 25 mg twice daily and 10 and 25 mg once daily- with eight healthy subjects per panel. Within each panel, subjects were randomized (3:1) to oral apixaban or placebo for 7 days. Subjects underwent safety assessments and were monitored for adverse events (AEs). Blood samples were taken to measure apixaban plasma concentration, international normalized ratio (INR), activated partial thromboplastin time (aPTT) and modified prothrombin time (mPT). Forty-eight subjects were randomized and treated (apixaban, n = 36; placebo, n = 12); one subject receiving 2.5 mg twice daily discontinued due to AEs (headache and nausea). No dose limiting AEs were observed. Apixaban maximum plasma concentration was achieved ~3 h post-dose. Exposure increased approximately in proportion to dose. Apixaban steady-state concentrations were reached by day 3, with an accumulation index of 1.3-1.9. Peak : trough ratios were lower for twice daily vs. once daily regimens. Clotting times showed dose-related increases tracking the plasma concentration-time profile. Multiple oral doses of apixaban were safe and well tolerated over a 10-fold dose range, with pharmacokinetics with low variability and concentration-related increases in clotting time measures.

A study to assess the pharmacokinetics (PK) of a single infusion of cetuximab (IMC-C225)

3084 Background: Cetuximab is a chimeric monoclonal antibody that selectively binds to the Epidermal Growth Factor Receptor (EGFR). This study examined single dose PK of cetuximab across a 10-fold dose range (50–500mg/m2). Methods: Thirty-nine patients with refractory solid tumors of epithelial origin were randomized to a single IV infusion of cetuximab of 50, 100, 250, 400 or 500 mg/m2. Blood samples for PK analysis were collected for up to three weeks following the first cetuximab dose. After 22 days, all patients were then administered weekly dosing of cetuximab at 250 mg/m2. Results: Results from noncompartmental PK analysis are summarized in the Table. Cetuximab serum concentrations reached a maximum at approximately 3h after initiation of infusion and then declined slowly thereafter. Concentrations approached baseline levels within 504 h for the majority of patients. Mean Cmax values for Cetuximab increased in a slightly greater than dose-proportional manner up to 500 mg/m2 while mean AUCinf values increased in a greater than dose-proportional manner up to 500 mg/m2. Cetuximab monotherapy was well tolerated. Approximately 60% of the patients experienced a skin rash, the majority of which was of Grade 1–2 in severity. Although no partial responses were seen, approximately 30 % of the patients achieved stable disease. Conclusions: The pharmacokinetic behavior of cetuximab demonstrated in this study is consistent with findings from previous studies. The measured half-life at the 250 mg/m2 dose of 71 hours is supportive of weekly dosing. Cetuximab is rapidly cleared at lower doses; however, clearance appears saturable at doses greater than 250 mg/m2. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Bristol-Myers Squibb Vertex Bristol-Myers Squibb Bristol-Myers Squibb Bristol-Myers Squibb

Extent and timeliness of tissue repair determines the dose-related hepatotoxicity of chloroform.

As a part of mixture toxicity studies, the objective of the present investigation was to validate the hypothesis that the rate and extent of liver tissue repair response to a given dose determines the end result of toxicity (death or recovery), regardless of the mechanisms by which injury is inflicted, using a well-known environmental pollutant, chloroform (CHCl(3)). In future, the data will be used to compare with the results of mixtures containing CHCl(3) to aid in characterizing the safety of chemical mixtures and to construct a physiologically based pharmacokinetic (PBPK) model for dose, route, and species extrapolation. Hepatotoxicity and tissue repair were measured in male Sprague-Dawley rats (S-D) receiving a 10-fold dose range of CHCl(3) (74, 185, 370, and 740 mg/kg, IP) during a time course of 0 to 96 hours. Liver injury, as assessed by plasma alanine aminotransferase (ALT) and sorbitol dehydrogenase (SDH) elevation, increased with dose over the 10-fold dose range. Because CHCl(3) is also known to cause kidney damage, blood urea nitrogen (BUN) and creatinine were measured to evaluate the kidney injury. With doses up to 370 mg/kg, liver injury increased in a dose-related fashion, which peaked at 24 hours and returned to normal after 48 hours, whereas at highest dose (740 mg/kg), the injury was progressive resulting in 90% mortality. Blood and liver CHCl(3) levels were quantified using gas chromatography (GC) over a time course of 30 to 360 minutes. The dose-related increase in the blood and liver CHCl(3) levels were consistent with dose-dependent liver injury. Tissue regeneration response, as measured by [(3)H]-thymidine incorporation into hepatocellular nuclear DNA peaked at 36 hours in rats treated with the lower two doses of CHCl(3) (74 and 185 mg/kg). Further increase in CHCl(3) dose to 370 mg/kg resulted in an earlier increase in [(3)H]-thymidine incorporation at 24 hours, which peaked at 36 hours. However, at the highest dose of CHCl(3) (740 mg/kg), tissue repair was delayed and attenuated, allowing for unrestrained progression of liver injury. The kidney injury markers after CHCl(3) administration were not different from controls. These results support the concept that in addition to the magnitude of tissue repair response, the time at which this response occurs is critical in restraining the progression of injury. Measuring tissue repair and injury as simultaneous biological responses to toxic agents might increase the usefulness of dose-response paradigms in predictive toxicology and risk assessment. Although the dosimetry of the present study was well beyond the environmental exposure levels of CHCl(3), a PBPK model will be developed in future based upon these data to evaluate the effects at environmental levels.

Percutaneous Absorption and Disposition of [14C]Chlordecone in Young and Adult Female Rats

The objective of this study was to evaluate the effect of age and dosage on percutaneous absorption and disposition of [14C]chlordecone (Kepone) and to describe results using a physiological based pharmacokinetic (PBPK) model. Female Fischer 344 rats 33 and 82 days old were used as the young and adult animal models, respectively, and were studied over a 10-fold dose range. [14C]Chlordecone (0.286 μmol/cm2) was applied to dorsal skin (2.3% BSA) and radioactivity was quantified in selected tissues and excreta up to 120 h. Absorption and disposition were also determined at three dose levels up to 2.68 μmol/cm2; fraction absorbed decreased as dose increased.In vitropercutaneous absorption was measured by static and flow-through methods; these yielded similar penetration rates, which were lower than those obtainedin vivo.In vivopercutaneous absorption over 120 h was 14.4±0.99 and 14.2±1.5% dose in young and adults, respectively. Organ and tissue content increased over time (carcass>liver>kidney), indicating prolonged absorption. Statistical differences between young and old were found for liver, skin, and urine, but not for absorption. Excretion occurred primarily in feces, but also in urine. A biophysically based percutaneous model was fitted to both young and adultin vivoabsorption data. This was embedded in a whole body PBPK model which, upon optimization with SAAM II, estimated apparent tissue partition coefficients, urinary and fecal excretion rates, and parameters characterizing hepatic nonlinear uptake of bound chlordecone. The model reasonably predicted tissue chlordecone content at higher doses, when decreased absorption was accounted for.

Tissue Repair Response as a Function of Dose during Trichloroethylene Hepatotoxicity

Trichloroethylene (TCE), a widely used organic solvent and degreasing agent, is regarded as a hepatotoxicant. The objective of the present studies was to investigate whether the extent and timeliness of tissue repair has a determining influence on the ultimate outcome of hepatotoxicity. Male Sprague-Dawley rats (200-250 g) were injected with a 10-fold dose range of TCE and hepatotoxicity and tissue repair were studied during a time course of 0 to 96 h. Light microscopic changes as evaluated by H&E-stained liver sections revealed a dose-dependent necrosis of hepatic cells. Maximum liver cell necrosis was observed at 48 h after the TCE administration. However, liver injury as assessed by plasma sorbitol dehydrogenase (SDH) showed a dose response over a 10-fold dose range only at 6 h, whereas alanine aminotransferase (ALT) did not show a dose response at any of the time points studied. A low dose of TCE (250 mg/kg) showed an increase in SDH at all time points up to 96 h without peak levels, whereas higher doses showed peak only at 6 h. At later time points SDH declined but remained above normal. In vitro addition of trichloroacetic acid, a metabolite of TCE to plasma, decreased the activities of SDH and ALT indicating that metabolites formed during TCE toxicity may interfere with plasma enzyme activities in vivo. This indicates that the lack of dose-related increase in SDH and ALT activities may be because of interference by the TCE metabolite. Tissue regeneration response as measured by [3H]thymidine incorporation into hepatocellular nuclear DNA was stimulated maximally at 24 h after 500 mg/kg TCE administration. A higher dose of TCE led to a delay and diminishment in [3H]thymidine incorporation. At a low dose of TCE (250 mg/kg) [3H]thymidine incorporation peaked at 48 h and this could be attributed to very low or minimal injury caused by this dose. With higher doses tissue repair was delayed and attenuated allowing for unrestrained progression of liver injury. These results support the concept that the toxicity and repair are opposing responses and that a dose-related increase in tissue repair represents a dynamic, quantifiable compensatory mechanism.

Comparative metabolism of high doses of aspirin in man and rat

1. Metabolism of aspirin was studied in 10 human volunteers who took a therapeutic dose (600 mg) by mouth and in nine patients who took aspirin in overdose. 2. Salicyluric acid was the major urinary metabolite in volunteers (63.1 +/- 8.4% of dose in 0-8 h). In overdose patients, salicyluric acid in urine was decreased (30.0 +/- 8.2%, 0-24 h, P less than 0.001) and there was increased elimination of salicyclic acid (34.1%, P less than 0.005), salicyl acyl glucuronide (14.4%, P less than 0.05) and gentisuric acid (5.3%). 3. Metabolism of orally administered 14C-aspirin in rats over a 10-fold dose range (10-100 mg/kg) resulted in excretion of 81-91% dose in urine in the first 24 h. Salicyclic acid was the major urinary metabolite (43-51% dose). Excretion of salicyluric acid decreased with increasing dose, whereas gentisic acid and salicyl phenolic and acyl glucuronides increased. 4. The profile of aspirin metabolites was qualitatively similar in man and rat but there were quantitative differences. Limited capacity to form salicyluric acid was observed in both species. Dependence on this pathway in rat was low and was compensated by increased utilization of other routes; dependence on salicyluric acid formation in man was high and in overdose, compensation by other routes was incomplete.

Oral absorption of 21-Corticosteroid Esters: A Function of Aqueous Stability and Intestinal Enzyme Activity and Distribution

The intestinal absorption of hydrocortisone and prednisolone are compared with three water-soluble derivatives (succinate, phosphate, and lysinate) in experiments at two levels of biological system complexity. Rates of absorption are compared by measuring permeabilities from rat intestinal perfusions of drugs and derivatives in solution. Extents of absorption are compared over a 10-fold dose range of parent steroid and with the steroid derivatives by measuring plasma levels from solid oral dosage in dogs. While the parent steroids are well absorbed over the entire length of the intestinal tract, variability in plasma levels is observed at higher doses. Limited solubility and resultant dissolution rate variability are likely to be playing a role in the early erratic blood level profiles found at higher doses. While the soluble prodrugs have a dissolution rate advantage which results in a greater concentration gradient, their absorption is limited by their aqueous luminal stability, their polarity and resultant passive membrane permeability, and the distribution and activity of enzyme reconversion sites in the intestinal tract. The unstable lysinate ester, targeted for aminopeptidase, has an absorption profile and permeability similar to that of the parent steroid. The absorption of the moderately stable succinate ester is limited by its polarity and the activity of intestinal esterases. The stable phosphate derivative is well absorbed in the upper intestine, where high levels of alkaline phosphatase exist, while the prodrug polarity and dropoff of enzyme activity limit its absorption from the lower gastrointestinal (Gl) tract.

The carcinogenicity of 15,16-dihydro-11-methyl-cyclopenta[a]phenanthren-17-one

Direct comparison of skin-tumour induction by 15,16-dihydro-11-methylcyclopenta[a]phenanthren-17-one (I) and by benzo[a]pyrene on mouse skin, both by repeated application or by initiation with a single dose followed by promotion with croton oil, demonstrated that these two carcinogens have similar potency. After repeated application of (I) the mean latent period for skin-tumour induction was linearly related to the logarithm of the dose over a 10-fold dose range. Under these conditions, application of the aryl-hydrocarbon-hydroxylase inhibitor 7,8-benzoflavone together with (I) inhibited tumour induction by about 40%. By contrast, in the 2-stage experiment, little effect on tumour incidence or latent period was observed when this inhibitor was applied with the single initiating dose of (I). Co-administration of the epoxide-hydratase inhibitor 1,1,1-trichloropropene oxide caused enhancement by shortening the latent period. After s.c. injection of (I) into mice, a similar number of tumours was induced on skin remote from the site of injection by promotion with corton oil begun either one week or 6 months after initiation. Gastric instillation of (I) into female rats induced mammary adenocarcinomas.