High-content, targeted RNA-seq screening in organoids for drug discovery in colorectal cancer.

Organoids allow the recapitulation of intestinal homeostasis and cancerogenesis in vitro; however, RNA sequencing (RNA-seq)-based methods for drug screens are missing. We develop targeted organoid sequencing (TORNADO-seq), a high-throughput, high-content drug discovery platform that uses targeted RNA-seq to monitor the expression of large gene signatures for the detailed evaluation of cellular phenotypes in organoids. TORNADO-seq is a fast, highly reproducible time- and cost-effective ($5 per sample) method that can probe cell mixtures and their differentiation state in the intestinal system. We apply this method to isolate drugs that enrich for differentiated cell phenotypes and show that these drugs are highly efficacious against cancer compared to wild-type organoids. Furthermore, TORNADO-seq facilitates in-depth insight into the mode of action of these drugs. Our technology can easily be adapted to many other systems and will allow for more systematic, large-scale, and quantitative approaches to study the biology of complex cellular systems.

Machine learning identifies candidates for drug repurposing in Alzheimer's disease.

Clinical trials of novel therapeutics for Alzheimer's Disease (AD) have consumed a large amount of time and resources with largely negative results. Repurposing drugs already approved by the Food and Drug Administration (FDA) for another indication is a more rapid and less expensive option. We present DRIAD (Drug Repurposing In AD), a machine learning framework that quantifies potential associations between the pathology of AD severity (the Braak stage) and molecular mechanisms as encoded in lists of gene names. DRIAD is applied to lists of genes arising from perturbations in differentiated human neural cell cultures by 80 FDA-approved and clinically tested drugs, producing a ranked list of possible repurposing candidates. Top-scoring drugs are inspected for common trends among their targets. We propose that the DRIAD method can be used to nominate drugs that, after additional validation and identification of relevant pharmacodynamic biomarker(s), could be readily evaluated in a clinical trial.

[Relationship between the number of prescribed medications and oral dysfunction in elderly individuals].

Objective　Polypharmacy in elderly individuals may cause reduced flow of saliva and xerostomia. A dry mouth can lead to poor oral function; however, there are no reports on the relationship between polypharmacy and subjective or objective oral dysfunction. The purpose of this study was to clarify the relationship between the number of prescribed medications and subjective and objective oral dysfunction.Methods　The subjects of this study were 215 community-dwelling, elderly individuals, aged 75 years or older, who visited the dental clinic in the Chubu region for a dental health examination from January to February 2019. A medical interview was conducted to assess three items that were related to subjective oral function and record four measurements related to objective oral function. In addition, information was collected on the diseases being treated and prescribed medications. A subject with a decrease in any of the three subjective oral function categories was considered to have subjective oral dysfunction. Objective oral dysfunction was analyzed with respect to two types of oral dysfunction: a decrease in all four objective oral functions and a decrease in two or more of the four objective oral functions. Logistic regression analysis was performed to examine the relationship between subjective and objective oral dysfunction after adjustment for sex, age group, inveterate disease, and the number of prescribed medications.Results　Individuals who had eight or more prescribed medications had lower subjective oral function than those with seven or fewer medications (odds ratio, 95% confidence interval: 2.3, 1.0-5.1; P<0.05). Individuals with eight or more medications had lower scores in all four objective oral functions than those with seven or fewer medications (4.4 : 1.5-12.6, P<0.01). A decrease in two or more of the four objective oral functions was related to 10 or more prescribed medications (4.3 : 1.2-16.2, P<0.05).　In addition, taking eight or more prescribed medications was associated with a decrease in either subjective oral function or all four objective oral functions (8.1 : 2.1-30.8, P<0.01). A decrease in either subjective oral function or two or more objective oral functions was related to taking 10 or more prescribed medications (4.9 : 1.6-15.6, P<0.01).Conclusion　In conclusion, more than eight prescribed pharmaceutical medications in the elderly is associated with subjective or objective oral dysfunction.

Inhibition of the ß-carbonic anhydrase from the protozoan pathogen Trichomonas vaginalis with sulphonamides.

Sulphonamides and their isosteres are classical inhibitors of the carbonic anhydrase (CAs, EC 4.2.1.1) metalloenzymes. The protozoan pathogen Trichomonas vaginalis encodes two such enzymes belonging to the ß-class, TvaCA1 and TvaCA2. Here we report the first sulphonamide inhibition study of TvaCA1, with a series of simple aromatic/heterocyclic primary sulphonamides as well as with clinically approved/investigational drugs for a range of pathologies (diuretics, antiglaucoma, antiepileptic, antiobesity, and antitumor drugs). TvaCA1 was effectively inhibited by acetazolamide and ethoxzolamide, with KIs of 391 and 283 nM, respectively, whereas many other simple or clinically used sulphonamides were micromolar inhibitors or did not efficiently inhibit the enzyme. Finding more effective TvaCA1 inhibitors may constitute an innovative approach for fighting trichomoniasis, a sexually transmitted infection, caused by T. vaginalis.

ConjuPepDB: a database of peptide-drug conjugates.

Peptide-drug conjugates are organic molecules composed of (i) a small drug molecule, (ii) a peptide and (iii) a linker. The drug molecule is mandatory for the biological action, however, its efficacy can be enhanced by targeted delivery, which often also reduces unwanted side effects. For site-specificity the peptide part is mainly responsible. The linker attaches chemically the drug to the peptide, but it could also be biodegradable which ensures controlled liberation of the small drug. Despite the importance of the field, there is no public comprehensive database on these species. Herein we describe ConjuPepBD, a freely available, fully annotated and manually curated database of peptide drug conjugates. ConjuPepDB contains basic information about the entries, e.g. CAS number. Furthermore, it also implies their biomedical application and the type of chemical conjugation employed. It covers more than 1600 conjugates from ∼230 publications. The web-interface is user-friendly, intuitive, and useable on several devices, e.g. phones, tablets, PCs. The webpage allows the user to search for content using numerous criteria, chemical structure and a help page is also provided. Besides giving quick insight for newcomers, ConjuPepDB is hoped to be also helpful for researchers from various related fields. The database is accessible at: https://conjupepdb.ttk.hu/.

Student Pharmacists' Ability to Organize Complex Medication Regimens According to the Universal Medication Schedule.

Objective. To assess student pharmacists' ability to impact the administration of complex prescription regimens using the universal medication schedule in a standardized laboratory exercise. Methods. First and third professional year (P1 and P3) student pharmacists at three colleges of pharmacy completed a required activity to simplify and organize a complex medication regimen. Using a medication box, students planned how and when they would advise a patient to take seven fictitious medications over a 24-hour period. Picture documentation of each students' activity was used for data analysis. Descriptive statistics were used to compare P1 and P3 students' performance, and an independent t test was used to assess the frequency of daily dosing. A chi-square analysis was used to compare differences between P1 and P3 students, and analysis of variance was used to compare differences among individual institutions. Results. Of 842 students invited, 459 P1 and 372 P3 students (98.7%) consented to participate. Student pharmacists recommended 5.1 (SD=1.0; Range=3-11) dosing intervals per 24 hours, with 27% of students successfully reducing the regimen to four total intervals. The P3 students were more effective than the P1 students in planning the number of dosing intervals (4.9 vs 5.4 per 24 hours). Conclusion. Student pharmacists may become more effective at organizing complex medication regimens as they proceed through the pharmacy curriculum and gain experience. Student pharmacists can translate what they learned from this exercise to potentially improve patients' self-organized medication regimens.

Physicochemical Properties, Biotransformation, and Transport Pathways of Established and Newly Approved Medications: A Systematic Review of the Top 200 Most Prescribed Drugs vs. the FDA-Approved Drugs Between 2005 and 2016.

BACKGROUND: Enzyme-mediated biotransformation of pharmacological agents is a crucial step in xenobiotic detoxification and drug disposition. Herein, we investigated the metabolism and physicochemical properties of the top 200 most prescribed drugs (established) as well as drugs approved by the US Food and Drug Administration (FDA) between 2005 and 2016 (newly approved). OBJECTIVE: Our objective was to capture the changing trends in the routes of administration, physicochemical properties, and prodrug medications, as well as the contributions of drug-metabolizing enzymes and transporters to drug clearance. METHODS: The University of Washington Drug Interaction Database (DIDB®) as well as other online resources (e.g., CenterWatch.com, Drugs.com, DrugBank.ca, and PubChem.ncbi.nlm.nih.gov) was used to collect and stratify the dataset required for exploring the above-mentioned trends. RESULTS: Analyses revealed that ~ 90% of all drugs in the established and newly approved drug lists were administered systemically (oral or intravenous). Meanwhile, the portion of biologics (molecular weight > 1 kDa) was 15 times greater in the newly approved list than established drugs. Additionally, there was a 4.5-fold increase in the number of compounds with a high calculated partition coefficient (cLogP > 3) and a high total polar surface area (> 75 Å2) in the newly approved drug vs. the established category. Further, prodrugs in established or newly approved lists were found to be converted to active compounds via hydrolysis, demethylases, and kinases. The contribution of cytochrome P450 (CYP) 3A4, as the major biotransformation pathway, has increased from 40% in the established drug list to 64% in the newly approved drug list. Moreover, the role of CYP1A2, CYP2C19, and CYP2D6 were decreased as major metabolizing enzymes among the newly approved medications. Among non-CYP major metabolizers, the contribution of alcohol dehydrogenases/aldehyde dehydrogenases (ADH/ALDH) and sulfotransferases decreased in the newly approved drugs compared with the established list. Furthermore, the highest contribution among uptake and efflux transporters was found for Organic Anion Transporting Polypeptide 1B1 (OATP1B1) and P-glycoprotein (P-gp), respectively. CONCLUSIONS: The higher portion of biologics in the newly approved drugs compared with the established list confirmed the growing demands for protein- and antibody-based therapies. Moreover, the larger number of hydrophilic drugs found in the newly approved list suggests that the probability of toxicity is likely to decrease. With regard to CYP-mediated major metabolism, CYP3A5 showed an increased involvement owing to the identification of unique probe substrates to differentiate CYP3As. Furthermore, the contribution of OATP1B1 and P-gp did not show a significant shift in the newly approved drugs as compared to the established list because of their broad substrate specificity.

Exposure to phthalate-containing prescription drugs and the risk of colorectal adenocarcinoma: A Danish nationwide case-control study.

PURPOSE: Some drug products contain phthalates as excipients, and in vitro studies have demonstrated that phthalates interfere with cellular mechanisms involved in colorectal cancer development. We therefore examined the association between cumulative phthalate exposure from drug products and risk of colorectal adenocarcinomas. METHODS: We used the Danish Cancer Registry to identify all patients with incident colorectal adenocarcinoma from 2008 to 2015 (n = 25 814). Each cancer case was matched to ten population controls. Linking information from Danish registers, we quantified cumulative phthalate exposure to the ortho-phthalates diethyl phthalate (DEP) and dibutyl phthalate (DBP) as well as enteric phthalate polymers from orally administered drugs. The association between cumulative phthalate exposure and colorectal cancer was estimated using conditional logistic regression. RESULTS: Cumulative exposure to ortho-phthalates exceeding 500 mg was associated with lower odds of colorectal cancer diagnosis (ORadj  = 0.89; 95% CI, 0.81-0.96). Similar associations were observed for all DEP exposure exceeding 500 mg. Subgroup analysis excluding NSAID users, demonstrated that ortho-phthalate exposure was positively associated with colorectal cancer (ORadj  = 1.26; 95% CI, 1.05-1.51). CONCLUSION: We found an apparent overall protective effect of cumulative phthalate exposure from drug excipients for colorectal adenocarcinoma. Omitting NSAID users reversed the signal and suggested a slightly increased risk associated with high cumulative ortho-phthalate exposure.

The toxicological analysis of lead and cadmium in prescription food for special medical purposes and modified milk products for newborns and infants available in Polish pharmacies.

There is currently a lack of risk assessments around Pb and Cd in prescription food for special medical purposes (FSMPs) and modified milk products available in from pharmacies in Poland. This article describes and evaluates a toxicological analysis of lead and cadmium in prescription FSMPs (n = 6) and modified milk products (n = 6) available in Polish pharmacies. The metals were determined using electrothermal atomisation atomic absorption spectrometry. To make the appropriate risk assessment, we considered (1) the levels of lead and cadmium in a powdered product, (2) the amount of the metals consumed in one portion (single exposure), (3) the daily dose depending on age (months) and body weight and (4) the weekly intake of Pb and Cd in samples in comparison to provisional tolerable weekly intake (PTWI). The results demonstrated that the samples analysed do not represent a health hazard to newborns and infants. This study is important because of the concerns around the complex risk assessment of prescription FSMP's and modified milk products available in Polish pharmacies related to lead and cadmium.

Evaluation of a Food and Drug Administration Mandate to Limit Acetaminophen in Prescription Combination Products.

INTRODUCTION: In 2014, the US Food and Drug Administration limited the production of prescription acetaminophen-opioid combination products to 325 mg per dose unit. The goal of this mandate was to decrease the likelihood of unintentional acetaminophen hepatotoxicity. This study was designed to determine if this federal regulation has succeeded in reducing unintentional acetaminophen-induced hepatotoxicity from opioid combination products. METHODS: Using data from the National Poison Data System (NPDS), we analyzed all calls to US Poison Control Centers in the years 2013 and 2015 for acetaminophen-opioid combination product exposures. We then excluded cases that were classified as intentional and those aged 12 years and younger. We used a primary endpoint of N-acetylcysteine administration; secondary endpoints included evidence of hepatotoxicity as aspartate aminotransferase elevation, opioid antagonist administration and severity of overall medical outcome. RESULTS: A total of 18,259 calls between the two yearlong periods met inclusion criteria. 5.16 and 5.01% of calls resulted in N-acetylcysteine administration in 2013 and 2015, respectively. 3.63 and 4.02% received naloxone in 2013 and 2015, respectively, and 0.9% in each year developed hepatotoxicity. Rates of N-acetylcysteine administration, naloxone administration, and hepatotoxicity did not differ significantly between 2013 and 2015. Severity of medical outcome was worse in 2015 as compared to 2013 with more cases being categorized as "major effect" and fewer cases being categorized as "no effect." CONCLUSIONS: The Food and Drug Administration limitation on acetaminophen content per dose unit in opioid combination products did not reduce the occurrence of unintentional acetaminophen-induced hepatotoxicity or N-acetylcysteine administration as reported to NPDS.

Repurposing FDA-approved drugs for anti-aging therapies.

There is great interest in drugs that are capable of modulating multiple aging pathways, thereby delaying the onset and progression of aging. Effective strategies for drug development include the repurposing of existing drugs already approved by the FDA for human therapy. FDA approved drugs have known mechanisms of action and have been thoroughly screened for safety. Although there has been extensive scientific activity in repurposing drugs for disease therapy, there has been little testing of these drugs for their effects on aging. The pool of FDA approved drugs therefore represents a large reservoir of drug candidates with substantial potential for anti-aging therapy. In this paper we employ FINDSITEcomb, a powerful ligand homology modeling program, to identify binding partners for proteins produced by temperature sensing genes that have been implicated in aging. This list of drugs with potential to modulate aging rates was then tested experimentally for lifespan and healthspan extension using a small invertebrate model. Three protein targets of the rotifer Brachionus manjavacas corresponding to products of the transient receptor potential gene 7, ribosomal protein S6 polypeptide 2 gene, or forkhead box C gene, were screened against a compound library consisting of DrugBank drugs including 1347 FDA approved, non-nutraceutical molecules. Twenty nine drugs ranked in the top 1 % for binding to each target were subsequently included in our experimental analysis. Continuous exposure of rotifers to 1 µM naproxen significantly extended rotifer mean lifespan by 14 %. We used three endpoints to estimate rotifer health: swimming speed (mobility proxy), reproduction (overall vitality), and mitochondria activity (cellular senescence proxy). The natural decline in swimming speed with aging was more gradual when rotifers were exposed to three drugs, so that on day 6, mean swimming speed of females was 1.19 mm/s for naproxen (P = 0.038), 1.20 for fludarabine (P = 0.040), 1.35 for hydralazine (P = 0.038), as compared to 0.88 mm/s in the control. The average reproduction of control females in the second half of their reproductive lifespan was 1.08 per day. In contrast, females treated with 1 µM naproxen produced 1.4 offspring per day (P = 0.027) and females treated with 10 µM fludarabine or 1 µM hydralazine produced 1.72 (P = <0.001) and 1.66 (P = 0.001) offspring per day, respectively. Mitochondrial activity naturally declines with rotifer aging, but B. manjavacas treated with 1 µM hydralazine or 10 µM fludarabine retained 49 % (P = 0.038) and 89 % (P = 0.002) greater mitochondria activity, respectively, than untreated controls. Our results demonstrate that coupling computation to experimentation can quickly identify new drug candidates with anti-aging potential. Screening drugs for anti-aging effects using a rotifer bioassay is a powerful first step in identifying compounds worthy of follow-up in vertebrate models. Even if lifespan extension is not observed, certain drugs could improve healthspan, slowing age-dependent losses in mobility and vitality.

Application of the 4D fingerprint method with a robust scoring function for scaffold-hopping and drug repurposing strategies.

Two factors contribute to the inefficiency associated with screening pharmaceutical library collections as a means of identifying new drugs: [1] the limited success of virtual screening (VS) methods in identifying new scaffolds; [2] the limited accuracy of computational methods in predicting off-target effects. We recently introduced a 3D shape-based similarity algorithm of the SABRE program, which encodes a consensus molecular shape pattern of a set of active ligands into a 4D fingerprint descriptor. Here, we report a mathematical model for shape similarity comparisons and ligand database filtering using this 4D fingerprint method and benchmarked the scoring function HWK (Hamza-Wei-Korotkov), using the 81 targets of the DEKOIS database. Subsequently, we applied our combined 4D fingerprint and HWK scoring function VS approach in scaffold-hopping and drug repurposing using the National Cancer Institute (NCI) and Food and Drug Administration (FDA) databases, and we identified new inhibitors with different scaffolds of MycP1 protease from the mycobacterial ESX-1 secretion system. Experimental evaluation of nine compounds from the NCI database and three from the FDA database displayed IC50 values ranging from 70 to 100 µM against MycP1 and possessed high structural diversity, which provides departure points for further structure-activity relationship (SAR) optimization. In addition, this study demonstrates that the combination of our 4D fingerprint algorithm and the HWK scoring function may provide a means for identifying repurposed drugs for the treatment of infectious diseases and may be used in the drug-target profile strategy.

Prediction of polypharmacological profiles of drugs by the integration of chemical, side effect, and therapeutic space.

Prediction of polypharmacological profiles of drugs enables us to investigate drug side effects and further find their new indications, i.e. drug repositioning, which could reduce the costs while increase the productivity of drug discovery. Here we describe a new computational framework to predict polypharmacological profiles of drugs by the integration of chemical, side effect, and therapeutic space. On the basis of our previous developed drug side effects database, named MetaADEDB, a drug side effect similarity inference (DSESI) method was developed for drug-target interaction (DTI) prediction on a known DTI network connecting 621 approved drugs and 893 target proteins. The area under the receiver operating characteristic curve was 0.882 ± 0.011 averaged from 100 simulated tests of 10-fold cross-validation for the DSESI method, which is comparative with drug structural similarity inference and drug therapeutic similarity inference methods. Seven new predicted candidate target proteins for seven approved drugs were confirmed by published experiments, with the successful hit rate more than 15.9%. Moreover, network visualization of drug-target interactions and off-target side effect associations provide new mechanism-of-action of three approved antipsychotic drugs in a case study. The results indicated that the proposed methods could be helpful for prediction of polypharmacological profiles of drugs.

FDA-approved drugs selected using virtual screening bind specifically to G-quadruplex DNA.

Guanine-rich sequences found in telomeres and oncogene promoters have the ability to form G-quadruplex structures. In this paper we describe the use of a virtual screening assay to search a database of FDA-approved compounds for compounds with the potential to bind G-quadruplex DNA. More than 750 telomerase inhibitors were identified in a literature search as acting through G-quadruplex stabilization, and from evaluation of these compounds, theoretical models capable of discriminating new compounds that bind G-quadruplex DNA were developed. Six compounds predicted to bind to the G-quadruplex structure were tested for their ability to bind to the human telomeric DNA sequence. Prochloroperazine, promazine, and chlorpromazine stabilized the G-quadruplex structure as determined by fluorescence resonance energy transfer techniques. These compounds also bound to promoter sequences of oncogenes such as c-myc and K-ras. Amitriptyline, imipramine, and loxapine were less stabilizing but did bind to the G-quadruplex. The ability of prochloroperazine, promazine, and chlorpromazine to recognize G-quadruplex structures was confirmed using a fluorescent intercalator displacement assay, in which displacement of thiazole orange from G-quadruplex structures was demonstrated. Interestingly, these compounds exhibited selectivity for the G-quadruplex structure as all had poor affinity for the duplex sequence.

Amphiphilic cylindrical copolypeptide brushes as potential nanocarriers for the simultaneous encapsulation of hydrophobic and cationic drugs.

Cylindrical copolypeptide brushes PLLF-g-(PLF-b-PLG) with poly(L-lysine-co-L-phenylalanine) (PLLF) as the backbone and poly(L-phenylalanine)-b-poly(L-glutamic acid) (PLF-b-PLG) as the side chains have been synthesized and evaluated as drug delivery carriers. The synthesized copolypeptide brushes were characterized by (1)H NMR, gel permeation chromatography (GPC), and transmission electron microscopy (TEM). In aqueous solution, the copolypeptide brushes adopt cylindrical morphologies and resemble unimolecular polymeric micelles with a hydrophobic poly(L-phenylalanine) core and a hydrophilic poly(L-glutamate) shell. An encapsulation study demonstrated that these water soluble, biodegradable copolypeptide brushes encapsulate hydrophobic compounds and cationic hydrophilic guest molecules simultaneously. Furthermore, the encapsulated cationic model compounds exhibit a pH-responsive releasing property.

Drug effect prediction by polypharmacology-based interaction profiling.

Most drugs exert their effects via multitarget interactions, as hypothesized by polypharmacology. While these multitarget interactions are responsible for the clinical effect profiles of drugs, current methods have failed to uncover the complex relationships between them. Here, we introduce an approach which is able to relate complex drug-protein interaction profiles with effect profiles. Structural data and registered effect profiles of all small-molecule drugs were collected, and interactions to a series of nontarget protein binding sites of each drug were calculated. Statistical analyses confirmed a close relationship between the studied 177 major effect categories and interaction profiles of ca. 1200 FDA-approved small-molecule drugs. On the basis of this relationship, the effect profiles of drugs were revealed in their entirety, and hitherto uncovered effects could be predicted in a systematic manner. Our results show that the prediction power is independent of the composition of the protein set used for interaction profile generation.

Prediction of off-target effects on angiotensin-converting enzyme 2.

The authors describe a structure-based strategy to identify therapeutically beneficial off-target effects by screening a chemical library of Food and Drug Administration (FDA)-approved small-molecule drugs matching pharmacophores defined for specific target proteins. They applied this strategy to angiotensin-converting enzyme 2 (ACE2), an enzyme that generates vasodilatory peptides and promotes protection from hypertension-associated cardiovascular disease. The conformation-based structural selection method by molecular docking using DOCK allowed them to identify a series of FDA-approved drugs that enhance catalytic efficiency of ACE2 in vitro. These data demonstrate that libraries of approved drugs can be rapidly screened to identify potential side effects due to interactions with specific proteins other than the intended targets.

Dual inhibition: a novel promising pharmacological approach for different disease conditions.

To overcome the problems associated with polypharmacy, which include medication non compliance, adverse drug reactions, drug-drug interactions and increased pill-burden, various strategies, such as sustained-release drugs and fixed-dose combination regimens (polypills), have been developed. Out of these, a novel and very much promising approach is the use of dual-action drugs. Amongst the dual-action drugs, there is a class of compounds known as dual inhibitors, which possess the dual inhibitory activity. The most common examples of dual inhibitors are rivastigmine, ladostigil, asenapine, phenserine, amitriptyline, clomipramine, doxepin and desipramine. This review article focuses on the conventional drugs used in different diseases which possess dual inhibition activity as well as those which are still in the preclinical/clinical phase.

In vitro screening of 50 highly prescribed drugs for thiol adduct formation--comparison of potential for drug-induced toxicity and extent of adduct formation.

Reactive metabolite formation has been associated with drug-induced liver, skin, and hematopoietic toxicity of many drugs that has resulted in serious clinical toxicity, leading to clinical development failure, black box warnings, or, in some cases, withdrawal from the market. In vitro and in vivo screening for reactive metabolite formation has been proposed and widely adopted in the pharmaceutical industry with the aim of minimizing the property and thus the risk of drug-induced toxicity (DIT). One of the most common screening methods is in vitro thiol trapping of reactive metabolites. Although it is well-documented that many hepatotoxins form thiol adducts, there is no literature describing the adduct formation potential of safer drugs that are widely used. The objective of this study was to quantitatively assess the thiol adduct formation potential of 50 drugs (10 associated with DIT and 40 not associated) and document apparent differences in adduct formation between toxic and safer drugs. Dansyl glutathione was used as a trapping agent to aid the quantitation of adducts following in vitro incubation of drugs with human liver microsomes in the presence and absence of NADPH. Metabolic turnover of these drugs was also monitored by LC/UV. Overall, 15 out of the 50 drugs screened formed detectable levels of thiol adducts. There were general trends toward more positive findings in the DIT group vs the non-DIT group. These trends became more marked when the relative amount of thiol adducts was taken into account and improved further when dose and total daily reactive metabolite burdens were considered. In conclusion, there appears to be a general trend between the extent of thiol adduct formation and the potential for DIT, which would support the preclinical measurement and minimization of the property through screening of thiol adduct formation as part of an overall discovery optimization paradigm.