Design, synthesis, and preliminary evaluation of a potential synthetic opioid rescue agent.

BACKGROUND: One of the most prominent opioid analgesics in the United States is the high potency agonist fentanyl. It is used in the treatment of acute and chronic pain and as an anesthetic adjuvant. When used inappropriately, however, ingestion of just a few milligrams of fentanyl or other synthetic opioid can cause opioid-induced respiratory depression (OIRD), often leading to death. Currently, the treatment of choice for OIRD is the opioid receptor antagonist naloxone. Recent reports, however, suggest that higher doses or repeated dosing of naloxone (due to recurrence of respiratory depression) may be required to reverse fully fentanyl-induced respiratory depression, rendering this treatment inadequate. To combat this synthetic opioid overdose crisis, this research aims at identifying a novel opioid reversal agent with enhanced efficacy towards fentanyl and other synthetic opioids. METHODS: A series of naltrexone analogues were characterized for their ability to antagonize the effects of fentanyl in vitro utilizing a modified forskolin-induced cAMP accumulation assay. Lead analogue 29 was chosen to undergo further PK studies, followed by in vivo pharmacological analysis to determine its ability to antagonize opioid-induced antinociception in the hot plate assay. RESULTS: A series of potent MOR antagonists were identified, including the highly potent analogue 29 (IC50 = 2.06 nM). Follow-up PK studies revealed 29 to possess near 100% bioavailability following IP administration. Brain concentrations of 29 surpassed plasma concentrations, with an apparent terminal half-life of ~ 80 min in mice. In the hot plate assay, 29 dose-dependently (0.01-0.1 mg/kg; IP) and fully antagonized the antinociception induced by oxycodone (5.6 mg/kg; IP). Furthermore, the dose of 29 that is fully effective in preventing oxycodone-induced antinociception (0.1 mg/kg) was ineffective against locomotor deficits caused by the KOR agonist U50,488. CONCLUSIONS: Methods have been developed that have utility to identify enhanced rescue agents for the treatment of OIRD. Analogue 29, possessing potent MOR antagonist activity in vitro and in vivo, provides a promising lead in our search for an enhanced synthetic opioid rescue agent.

OleD Loki as a Catalyst for Hydroxamate Glycosylation.

Herein we describe the ability of the permissive glycosyltransferase (GT) OleD Loki to convert a diverse set of >15 histone deacetylase (HDAC) inhibitors (HDACis) into their corresponding hydroxamate glycosyl esters. Representative glycosyl esters were subsequently evaluated in assays for cancer cell line cytotoxicity, chemical and enzymatic stability, and axolotl embryo tail regeneration. Computational substrate docking models were predictive of enzyme-catalyzed turnover and suggest certain HDACis may form unproductive, potentially inhibitory, complexes with GTs.

GZ-11608, a Vesicular Monoamine Transporter-2 Inhibitor, Decreases the Neurochemical and Behavioral Effects of Methamphetamine.

Despite escalating methamphetamine use and high relapse rates, pharmacotherapeutics for methamphetamine use disorders are not available. Our iterative drug discovery program had found that R-N-(1,2-dihydroxypropyl)-2,6-cis-di-(4-methoxyphenethyl)piperidine hydrochloride (GZ-793A), a selective vesicular monoamine transporter-2 (VMAT2) inhibitor, specifically decreased methamphetamine's behavioral effects. However, GZ-793A inhibited human-ether-a-go-go-related gene (hERG) channels, suggesting cardiotoxicity and prohibiting clinical development. The current study determined if replacement of GZ-793A's piperidine ring with a phenylalkyl group to yield S-3-(4-methoxyphenyl)-N-(1-phenylpropan-2-yl)propan-1-amine (GZ-11608) diminished hERG interaction while retaining pharmacological efficacy. VMAT2 inhibition, target selectivity, and mechanism of GZ-11608-induced inhibition of methamphetamine-evoked vesicular dopamine release were determined. We used GZ-11608 doses that decreased methamphetamine-sensitized activity to evaluate the potential exacerbation of methamphetamine-induced dopaminergic neurotoxicity. GZ-11608-induced decreases in methamphetamine reinforcement and abuse liability were determined using self-administration, reinstatement, and substitution assays. Results show that GZ-11608 exhibited high affinity (Ki = 25 nM) and selectivity (92-1180-fold) for VMAT2 over nicotinic receptors, dopamine transporter, and hERG, suggesting low side-effects. GZ-11608 (EC50 = 620 nM) released vesicular dopamine 25-fold less potently than it inhibited VMAT2 dopamine uptake. GZ-11608 competitively inhibited methamphetamine-evoked vesicular dopamine release (Schild regression slope = 0.9 ± 0.13). GZ-11608 decreased methamphetamine sensitization without altering striatal dopamine content or exacerbating methamphetamine-induced dopamine depletion, revealing efficacy without neurotoxicity. GZ-11608 exhibited linear pharmacokinetics and rapid brain penetration. GZ-11608 decreased methamphetamine self-administration, and this effect was not surmounted by increasing methamphetamine unit doses. GZ-11608 reduced cue- and methamphetamine-induced reinstatement, suggesting potential to prevent relapse. GZ-11608 neither served as a reinforcer nor substituted for methamphetamine, suggesting low abuse liability. Thus, GZ-11608, a potent and selective VMAT2 inhibitor, shows promise as a therapeutic for methamphetamine use disorder. SIGNIFICANCE STATEMENT: GZ-11608 is a potent and selective vesicular monoamine transporter-2 inhibitor that decreases methamphetamine-induced dopamine release from isolated synaptic vesicles from brain dopaminergic neurons. Results from behavioral studies show that GZ-11608 specifically decreases methamphetamine-sensitized locomotor activity, methamphetamine self-administration, and reinstatement of methamphetamine-seeking behavior, without exhibiting abuse liability. Tolerance does not develop to the efficacy of GZ-11608 to decrease the behavioral effects of methamphetamine. In conclusion, GZ-11608 is an outstanding lead in our search for a therapeutic to treat methamphetamine use disorder.

Population pharmacokinetic analysis of AR-67, a lactone stable camptothecin analogue, in cancer patients with solid tumors.

Background AR-67 is a novel camptothecin analogue at early stages of drug development. The phase 1 clinical trial in cancer patients with solid tumors was completed and a population pharmacokinetic model (POP PK) was developed to facilitate further development of this investigational agent. Methods Pharmacokinetic data collected in the phase 1 clinical trial were utilized for the development of a population POP PK by implementing the non-linear mixed effects approach. Patient characteristics at study entry were evaluated as covariates in the model. Subjects (N = 26) were treated at nine dosage levels (1.2-12.4 mg/m2/day) on a daily × 5 schedule. Hematological toxicity data were modeled against exposure metrics. Results A two-compartment POP PK model best described the disposition of AR-67 by fitting a total of 328 PK observations from 25 subjects. Following covariate model selection, age remained as a significant covariate on central volume. The final model provided a good fit for the concentration versus time data and PK parameters were estimated with good precision. Clearance, inter-compartmental clearance, central volume and peripheral volume were estimated to be 32.2 L/h, 28.6 L/h, 6.83 L and 25.0 L, respectively. Finally, exposure-pharmacodynamic analysis using Emax models showed that plasma drug concentration versus time profiles are better predictors of AR-67-related hematologic toxicity were better predictors of leukopenia and thrombocytopenia, as compared to total dose. Conclusions A POP PK model was developed to characterize AR-67 pharmacokinetics and identified age as a significant covariate. Exposure PK metrics Cmax and AUC were shown to predict hematological toxicity. Further efforts to identify clinically relevant determinants of AR-67 disposition and effects in a larger patient population are warranted.

Bioanalytical method for quantitative determination of mithramycin analogs in mouse plasma by HPLC-QTOF.

Mithramycin (MTM) has potent anticancer activity, but severe toxicities restrict its clinical use. Semi-synthetic approaches have yielded novel MTM analogs with potentially lower toxicity and similar efficacy. In an effort to transition these analogs into in vivo models, a bioanalytical method was developed for their quantification in mouse plasma. Here we present the validation of the method for the quantitation of mithramycin SA-tryptophan (MTMSA-Trp) as well as the applicability of the methodology for assaying additional analogs, including MTM, mithramycin SK (MTMSK) and mithramycin SA-phenylalanine (MTMSA-Phe) with run times of 6 min. Assay linearity ranged from 5 to 100 ng/mL. Accuracies of calibration standards and quality control samples were within 15% of nominal with precision variability of <20%. MTMSA-Trp was stable for 30 days at -80°C and for at least three freeze-thaw cycles. Methanol (-80°C) extraction afforded 92% of MTMSA-Trp from plasma. Calibration curves for MTM and analogs were also linear from ≤5 to 100 ng/mL. This versatile method was used to quantitate MTM analogs in plasma samples collected during preclinical pharmacokinetic studies.

Validation of a HPLC/MS method for simultaneous quantification of clonidine, morphine and its metabolites in human plasma.

A high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous quantification of morphine, morphine's major metabolites morphine-3-glucuronide and morphine-6-glucuronide, and clonidine, to support the pharmacokinetic analysis of an ongoing double-blinded randomized clinical trial that compares the use of morphine and clonidine in infants diagnosed with neonatal abstinence syndrome. Plasma samples were processed by solid-phase extraction and separated on an Inertsil ODS-3 (4 µm) column using an 0.1% formic acid in water-0.1% formic acid in methanol gradient. Detection of the analytes was conducted in the positive multiple reaction monitoring mode. The range of quantitation was 1-1000 ng/mL for morphine, morphine-3-glucuronide and morphine-6-glucuronide, and 0.25-100 ng/mL for clonidine. Intra-day and inter-day accuracy and precision were ≤15% for all analytes across the quantitation range. Extraction recovery rates were ≥94% for morphine, ≥90% for M3G, ≥87% for M6G and ≥ 79% for clonidine. Matrix effect ranged from 85-94% for clonidine to 101-106% for M3G. The method fulfilled all predetermined acceptance criteria and required only 100 µL of starting plasma volume. Furthermore, it was successfully applied to 30 clinical trial plasma samples.

Pharmacokinetic modeling of the blood-stable camptothecin analog AR-67 in two different formulations.

AR-67 is a lipophilic camptothecin analog currently under clinical investigation using a Cremophor EL based formulation. However, as potential toxicity limitations exist in the clinical use of Cremophor, an alternative cyclodextrin (SBE-ß-CD) based formulation has been proposed. Pharmacokinetic (PK) studies were conducted in mice and the SBE-ß-CD based formulation was compared with the Cremophor EL formulation. PK studies were conducted following intravenous or oral administration of AR-67 in either Cremophor or SBE-ß-CD formulation in mice. Noncompartmental analysis was used to determine the plasma and tissue drug distribution. A non-linear mixed effects (population) PK model was developed to fit both the oral and intravenous data and to estimate key PK parameters. The effect of formulation was explored as a covariate in the PK model. AR-67 in the SBE-ß-CD formulation had similar plasma PK and biodistribution to that in the Cremophor EL formulation. The proposed two-compartment model described the plasma PK of AR-67 in both formulations adequately. AR-67 in the SBE-ß-CD formulation exhibited dose linearity following both oral and intravenous administration. Our studies indicate that SBE-ß-CD is a viable alternative to Cremophor EL as a pharmaceutical excipient for formulating AR-67.

Phase 1b trial of proteasome inhibitor carfilzomib with irinotecan in lung cancer and other irinotecan-sensitive malignancies that have progressed on prior therapy (Onyx IST reference number: CAR-IST-553).

Introduction Proteasome inhibition is an established therapy for many malignancies. Carfilzomib, a novel proteasome inhibitor, was combined with irinotecan to provide a synergistic approach in relapsed, irinotecan-sensitive cancers. Materials and Methods Patients with relapsed irinotecan-sensitive cancers received carfilzomib (Day 1, 2, 8, 9, 15, and 16) at three dose levels (20/27 mg/m2, 20/36 mg/m2 and 20/45 mg/m2/day) in combination with irinotecan (Days 1, 8 and 15) at 125 mg/m2/day. Key eligibility criteria included measurable disease, a Zubrod PS of 0 or 1, and acceptable organ function. Patients with stable asymptomatic brain metastases were eligible. Dose escalation utilized a standard 3 + 3 design. Results Overall, 16 patients were enrolled to three dose levels, with four patients replaced. Three patients experienced dose limiting toxicity (DLT) and the maximum tolerated dose (MTD) was exceeded in Cohort 3. The RP2 dose was carfilzomib 20/36 mg/m2 (given on Days 1, 2, 8, 9, 15, and 16) and irinotecan 125 mg/m2 (Days 1, 8 and 15). Common Grade (Gr) 3 and 4 toxicities included fatigue (19%), thrombocytopenia (19%), and diarrhea (13%). Conclusions Irinotecan and carfilzomib were well tolerated, with common toxicities of fatigue, thrombocytopenia and neutropenic fever. Objective clinical response was 19% (one confirmed partial response (PR) in small cell lung cancer (SCLC) and two unconfirmed); stable disease (SD) was 6% for a disease control rate (DCR) of 25%. The recommended phase II dose was carfilzomib 20/36 mg/m2 and irinotecan125 mg/m2. The phase II evaluation is ongoing in relapsed small cell lung cancer.

The combination of ezetimibe and ursodiol promotes fecal sterol excretion and reveals a G5G8-independent pathway for cholesterol elimination.

Previous studies suggest an interdependent relationship between liver and intestine for cholesterol elimination from the body. We hypothesized that a combination of ursodiol (Urso) and ezetimibe (EZ) could increase biliary secretion and reduce cholesterol reabsorption, respectively, to promote cholesterol excretion. Treatment with Urso increased hepatic ABCG5 ABCG8 (G5G8) protein and both biliary and fecal sterols in a dose-dependent manner. To determine whether the drug combination (Urso-EZ) further increased cholesterol excretion, mice were treated with Urso alone or in combination with two doses of EZ. EZ produced an additive and dose-dependent increase in fecal neutral sterol (FNS) elimination in the presence of Urso. Finally, we sequentially treated wide-type and G5G8-deficient mice with Urso and Urso-EZ to determine the extent to which these effects were G5G8 dependent. Although biliary and FNS were invariably lower in G5G8 KO mice, the relative increase in FNS following treatment with Urso alone or the Urso-EZ combination was not affected by genotype. In conclusion, Urso increases G5G8, biliary cholesterol secretion, and FNS and acts additively with EZ to promote fecal sterol excretion. However, the stimulatory effect of these agents was not G5G8 dependent.

Cytotoxic Indolocarbazoles from Actinomadura melliaura ATCC 39691.

Actinomadura melliaura ATCC 39691, a strain isolated from a soil sample collected in Bristol Cove, California, is a known producer of the disaccharide-substituted AT2433 indolocarbazoles (6-9). Reinvestigation of this strain using new media conditions led to >40-fold improvement in the production of previously reported AT2433 metabolites and the isolation and structure elucidation of the four new analogues, AT2433-A3, A4, A5, and B3 (1-4). The availability of this broader set of compounds enabled a subsequent small antibacterial/fungal/cancer SAR study that revealed disaccharyl substitution, N-6 methylation, and C-11 chlorination as key modulators of bioactivity. The slightly improved anticancer potency of the newly reported N-6-desmethyl 1 (compared to 6) contrasts extensive SAR of monoglycosylated rebeccamycin-type topoisomerase I inhibitors where N-6 alkylation has contributed to improved potency and ADME. Complete 2D NMR assignments for the known metabolite BMY-41219 (5) and (13)C NMR spectroscopic data for the known analogue AT2433-B1 (7) are also provided for the first time.

Enzymatic methylation and structure-activity-relationship studies on polycarcin V, a gilvocarcin-type antitumor agent.

Polycarcin V, a polyketide natural product of Streptomyces polyformus, was chosen to study structure-activity relationships of the gilvocarcin group of antitumor antibiotics due to a similar chemical structure and comparable bioactivity with gilvocarcin V, the principle compound of this group, and the feasibility of enzymatic modifications of its sugar moiety by auxiliary O-methyltransferases. Such enzymes were used to modify the interaction of the drug with histone H3, the biological target that interacts with the sugar moiety. Cytotoxicity assays revealed that a free 2'-OH group of the sugar moiety is essential to maintain the bioactivity of polycarcin V, apparently an important hydrogen bond donor for the interaction with histone H3, and converting 3'-OH into an OCH3 group improved the bioactivity. Bis-methylated polycarcin derivatives revealed weaker activity than the parent compound, indicating that at least two hydrogen bond donors in the sugar are necessary for optimal binding.

Neonatal Abstinence Signs during Treatment: Trajectory, Resurgence and Heterogeneity.

Neonatal abstinence syndrome (NAS) presents with a varying severity of withdrawal signs and length of treatment (LOT). We examined the course and relevance of each of the NAS withdrawal signs during treatment in a sample of 182 infants with any prenatal opioid exposure, gestational age ≥ 35 weeks, without other medical conditions, and meeting the criteria for pharmacological treatment. Infants were monitored using the Finnegan Neonatal Abstinence Scoring Tool. Daily mean Finnegan scores were estimated using linear mixed models with random subject effects to account for repeated withdrawal scores from the same subject. Daily item prevalence was estimated using generalized estimating equations with a within-subject exchangeable correlation structure. The median LOT was 12.86 days. The prevalence of withdrawal signs decreased from day one to day three of treatment. However, certain central nervous system (CNS) and gastrointestinal (GI) signs showed sporadic increases in prevalence notable around two weeks of treatment, accounting for increases in Finnegan scores that guided pharmacotherapy. We question whether the resurgence of signs with a prolonged LOT is mainly a consequence of opioid tolerance or withdrawal. Monitoring CNS and GI signs throughout treatment is crucial. Future studies directed to better understand this clinical phenomenon may lead to the refining of NAS pharmacotherapy and perhaps the discovery of treatment alternatives.

The effect of breast cancer resistance protein, multidrug resistant protein 1, and organic anion-transporting polypeptide 1B3 on the antitumor efficacy of the lipophilic camptothecin 7-t-butyldimethylsilyl-10-hydroxycamptothecin (AR-67) in vitro.

AR-67 (7-t-butyldimethylsilyl-10-hydroxycamptothecin) is a lipophilic camptothecin analog, currently under early stage clinical trials. Transporters are known to have an impact on the disposition of camptothecins and on the response to chemotherapeutics in general due to their expression in tumor tissues. Therefore, we investigated the interplay between the breast cancer resistance protein (BCRP), multidrug resistant protein 1 (MDR1), and organic anion-transporting polypeptide (OATP) 1B1/1B3 transporters and AR-67 and their impact on the toxicity profile of AR-67. Using cell lines expressing the aforementioned transporters, we showed that the lipophilic AR-67 lactone form is a substrate for efflux transporters BCRP and MDR1. Additionally, OATP1B1 and OATP1B3 facilitated the uptake of AR-67 carboxylate in SLCO1B1- and SLCO1B3-transfected cell systems compared with the mock-transfected ones. Notably, both BCRP and MDR1 conferred resistance to AR-67 lactone. Prompted by recent studies showing increased OATP1B3 expression in certain cancer types, we investigated the effect of OATP1B3 expression on cell viability after exposure to AR-67 carboxylate. OATP1B3-expressing cells had increased carboxylate uptake as compared with mock-transfected cells but were not sensitized because the intracellular amount of lactone was 50-fold higher than that of carboxylate and comparable between OATP1B3-expressing and OATP1B3-nonexpressing cells. In conclusion, BCRP- and MDR1-mediated efflux of AR-67 lactone confers resistance to AR-67, but OATP1B3-mediated uptake of the AR-67 carboxylate does not sensitize OATP1B3-expressing tumor cells.

High payload dual therapeutic-imaging nanocarriers for triggered tumor delivery.

The in vitro and in vivo characterization of an optimized formulation of nanoparticles (NPs) loaded with a high content of dexamethasone palmitate (DEX-P), a chemotherapeutic adjuvant that decreases interstitial fluid pressure in tumors, and (111) In, a signaling agent, is described. These NPs are uniform in size and composition. Single photon emission computed tomography imaging demonstrates significant tumor uptake of (111) In-labeled DEX-P NPs in tumor-bearing mice. As with many nanoparticle-based drug delivery systems, significant liver accumulation is observed. Assessment of liver histology and blood tests show no apparent hepatic or renal toxicity of the DEX-P NPs. Conversion of DEX-P to DEX occurs when DEX-P NPs are incubated with mouse plasma, human tumor homogenate and ascites from tumor bearing mice, but not with human plasma. This conversion is slower in plasma from Es1(e) ((-/-)) /SCID mice, a potential alternative animal model that better mimics humans; however, plasma from these mice are not completely devoid of esterase activity. The difference between blood and tumor esterase activity in humans facilitates the delivery of DEX-P NPs to tumors and the release of dexamethasone by an esterase trigger.

Pharmacologic properties of polyethylene glycol-modified Bacillus thiaminolyticus thiaminase I enzyme.

We have previously shown that the bacterial enzyme thiaminase 1 has antitumor activity. In an attempt to make thiaminase I a more effective pharmaceutical agent, we have modified it by adding polyethylene glycol (PEG) chains of various lengths. We were surprised to find that 5k-PEGylation eliminated thiaminase cytotoxic activity in all cell lines tested. Both native thiaminase and 5k-PEGylated thiaminase efficiently depleted thiamine from cell culture medium, and both could use intracellular phosphorylated thiamine as substrates. However, native enzyme more effectively depleted thiamine and thiamine diphosphate in RS4 leukemia cell cytosol, and native thiaminase depressed cellular respiration, whereas PEGylated thiaminase did not. Despite the lack of in vitro cytotoxicity, PEGylation markedly increased the in vivo toxicity of the enzyme. Pharmacokinetic studies revealed that the half-life of native thiaminase was 1.5 h compared with 34.4 h for the 5k-PEGylated enzyme. Serum thiamine levels were depleted by both native and 5k-PEGylated enzyme. Despite superior pharmacokinetics, 5k-PEGylated thiaminase showed no antitumor effect against an RS4 leukemia xenograft, in contrast to native thiaminase, which showed antitumor activity. PEGylation of thiaminase I has demonstrated that depression of mitochondrial function contributes, at least in part, to its anticancer activity. PEGylation also enhances plasma retention time, which increased its vivo toxicity and decreased its activity against a leukemia xenograft, the opposite of the desired effects. These studies suggest that the mechanism of anticancer cytotoxicity of thiaminase requires acute depression of cellular respiration, whereas systemic toxicity is related to the duration of extracellular thiamine depletion.

Stereoselective interaction of pantoprazole with ABCG2. II. In vitro flux analysis.

(-)Pantoprazole [(-)PAN] accumulated in rat milk stereoselectively, and this accumulation was attributed to rat Abcg2 (rAbcg2). In contrast, flux experiments at 25 µM showed that (+)pantoprazole [(+)PAN] was preferentially transported by rAbcg2. The purpose of the current study was to comprehensively evaluate the transport of PAN isomers in empty-Madin-Darby canine kidney II (MDCKII) and MDCKII cells expressing the human/rat (ABCG2/rAbcg2) isoforms at concentrations ranging from 3 to 200 µM. The apical-to-basolateral and basolateral-to-apical directional flux and the asymmetry efflux ratios were virtually identical for both isomers in empty (mock transfected)-MDCKII monolayers but were concentration dependent for both isomers in ABCG2 (human/rat)-MDCKII. Kinetic analysis using predicted cellular concentrations showed that (-)PAN had an 8-fold lower K(M) compared with (+)PAN for both rAbcg2 (0.25 versus 1.85 µM) and ABCG2 (0.6 versus 5.32 µM). (+)PAN had a 3-fold higher T(Max) compared with the (-)PAN for both rAbcg2 (7.86 versus 2.49 nmol/h · cm(2)) and ABCG2 (10.2 versus 3.29 nmol/h · cm(2)). Effective ABCG2 surface-area permeability of (-)PAN was 9920 and 5480 (µl/h)/cm(2) for rAbcg2 and ABCG2, respectively, compared with the (+)PAN isomer (4250 and 1920 µl/h · cm(2), respectively). These results indicate a stereoselective interaction of PAN with similar kinetic parameters for both human and rat ABCG2. (-)PAN is a better substrate than (+)PAN for ABCG2/rAbcg2 and provide a rationale for the preferential accumulation of (-)PAN into rat milk.

Pharmacokinetic modeling to assess factors affecting the oral bioavailability of the lactone and carboxylate forms of the lipophilic camptothecin analogue AR-67 in rats.

PURPOSE: Camptothecin analogues are anticancer drugs effective when dosed in protracted schedules. Such treatment is best suited for oral formulations. AR-67 is a novel lipophilic analogue with potent efficacy in preclinical models. Here we assessed factors that may influence its oral bioavailability in rats. METHODS: Plasma pharmacokinetic (PK) studies were conducted following administration of AR-67 lactone or carboxylate doses alone or after pre-dosing with inhibitors of the efflux transporters P-gp and Bcrp. A population PK model that simultaneously fitted to oral and intravenous data was used to estimate the bioavailability (F) and clearance of AR-67. RESULTS: An inverse Gaussian function was used as the oral input into the model and provided the best fits. Covariate analysis showed that the bioavailability of the lactone, but not its clearance, was dose dependent. Consistent with this observation, the bioavailability of AR-67 increased when animals were pretreated orally with GF120918 or Zosuquidar. CONCLUSION: Absorption of AR-67 is likely affected by solubility of its lactone form and interaction with efflux pumps in the gut. AR-67 appears to be absorbed as the lactone form, most likely due to gastric pH favoring its formation and predominance. F increased at higher doses suggesting saturation of efflux mechanisms.

Saquayamycins G-K, cytotoxic angucyclines from Streptomyces sp. Including two analogues bearing the aminosugar rednose.

Streptomyces sp. KY40-1, a strain isolated from the Kentucky Appalachian foothills, is the producer of moromycins A (18) and B (19). Further investigations of this strain led to the isolation and structure elucidation of the five new saquayamycins G-K (1-5), along with known compounds. Two of the new compounds bear the unusual aminosugar rednose, which was found here for the first time in angucyclines. The different attachment positions of this aminosugar in these two compounds indicate a high acceptor substrate flexibility of the responsible glycosyl transferase or alternatively the involvement of multiple glycosyl transferases. The cytotoxic activity of the isolated compounds was determined using human prostate cancer (PC-3) and non-small-cell lung cancer (H460) cell lines. Cell viability assays showed that saquayamycins J (4), K (5), A (7), and B (8) were most active in PC3 cells, with saquayamycin B (8) showing the highest activity (GI(50) = 0.0075 µM). The aminosugar-containing saquayamycins H (2) and saquayamycin B (8) showed the highest activity against H460 cells, with a GI(50) of 3.3 and 3.9 µM, respectively. The results presented here provide more insights into the structure-activity relationship of saquayamycins with respect to the nature, number, and linkage of sugar residues.

Assessment of the relative clinical utility of shortened Finnegan neonatal abstinence scoring tools.

OBJECTIVE: To assess the proposed shortened tools based on the Finnegan neonatal abstinence scoring tool (FNAS) for relative clinical utility. STUDY DESIGN: Retrospective study comparing shortened tools with FNAS on need for treatment, medication initiation cutoff score agreement, and length of treatment in 369 infants with prenatal opioid exposure using estimated areas under the receiver operating characteristic curves, Pearson and Spearman correlations, and proportion correctly classified, sensitivity, and specificity. RESULTS: The tools by Gomez et al. and Chervoneva et al. are most predictive of the FNAS cut-off values to initiate treatment, have cutoff values that best align with the FNAS cutoff values, and strongly correlate with the FNAS (r ≥ 0.88 corresponding to treatment initiation, r ≥ 0.83 during first 10 days of treatment). CONCLUSION: The tools of Gomez and Chervoneva demonstrated potential clinical usefulness by strongly associating with the need for treatment and monitoring the course of NAS therapy.

Metabolic pathways of the camptothecin analog AR-67.

7-tert-Butyldimethylsilyl-10-hydroxycamptothecin (AR-67; also known as DB-67) is a novel lipophilic camptothecin analog in early-phase anticancer clinical trials. In support of these studies, we evaluated the metabolism of AR-67 in vitro and identified potential metabolites in patient samples. The lactone form of AR-67 was found to be preferentially metabolized over AR-67 carboxylate in human microsomes. Subsequently, the lactone form was tested as a substrate in a panel of CYP450 and UDP-glucuronosyltransferase (UGT) enzymes known to metabolize the majority of clinically approved molecules. AR-67 was metabolized by CYP3A5, CYP3A4, CYP1A1, and CYP1A2, in order of activity. Extrahepatic UGT1A8 and UGT1A7 possessed at least 6-fold higher metabolizing activity than UGT1A1 and other UGT enzymes tested. CYP1A1 and UGT1A7 displayed Michaelis-Menten kinetics, whereas CYP3A4, CYP3A5, and UGT1A8 displayed kinetics consistent with substrate inhibition. Chromatographic analysis of representative patient plasma and urine samples demonstrated the presence of AR-67 glucuronides and oxidized products in the urine but only in very minimal amounts. We conclude that limited in vivo metabolism of AR-67 by UGT1A1 may partly explain the absence of AR-67 glucuronides in plasma and hypothesize that UGT1A8- and CYP3A-mediated biotransformation within the gastrointestinal epithelium may provide protective mechanisms against AR-67 gastrointestinal toxicity.