Screening for DNA adducts by data-dependent constant neutral loss-triple stage mass spectrometry with a linear quadrupole ion trap mass spectrometer.

International audienceA two-dimensional linear quadrupole ion trap mass spectrometer (LIT/MS) was employed to simultaneously screen for DNA adducts of environmental, dietary, and endogenous genotoxicants, by data-dependent constant neutral loss scanning followed by triple-stage mass spectrometry (CNL-MS3). The loss of the deoxyribose (dR) from the protonated DNA adducts ([M + H - 116]+) in the MS/MS scan mode triggered the acquisition of MS3 product ion spectra of the aglycone adducts [BH2]+. Five DNA adducts of the tobacco carcinogen 4-aminobiphenyl (4-ABP) were detected in human hepatocytes treated with 4-ABP, and three DNA adducts of the cooked-meat carcinogen 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) were identified in the livers of rats exposed to MeIQx, by the CNL-MS3 scan mode. Buccal cell DNA from tobacco smokers was screened for DNA adducts of various classes of carcinogens in tobacco smoke including 4-ABP, 2-amino-9H-pyrido[2,3-b]indole (AalphaC), and benzo[a]pyrene (BaP); the cooked-meat carcinogens MeIQx, AalphaC, and 2-amino-1-methyl-6-phenylmidazo[4,5-b]pyridine (PhIP); and the lipid peroxidation products acrolein (AC) and trans-4-hydroxynonenal (HNE). The CNL-MS3 scanning technique can be used to simultaneously screen for multiple DNA adducts derived from different classes of carcinogens, at levels of adduct modification approaching 1 adduct per 108 unmodified DNA bases, when 10 microg of DNA is employed for the assay

Cytochrome P450-Mediated Metabolism and DNA Binding of 2-Amino-1,7-dimethylimidazo[4,5-<i>g</i>]quinoxaline and Its Carcinogenic Isomer 2-Amino-3,8-dimethylimidazo[4,5-<i>f</i>]quinoxaline in Mice

2-Amino-1,7-dimethylimidazo­[4,5-<i>g</i>]­quinoxaline (MeI<i>g</i>Qx) is a recently discovered heterocyclic aromatic amine (HAA) that is formed during the cooking of meats. MeI<i>g</i>Qx is an isomer of 2-amino-3,8-dimethylmidazo­[4,5-<i>f</i>]­quinoxaline (MeIQx), a rodent carcinogen and possible human carcinogen that also occurs in cooked meats. MeI<i>g</i>Qx is a bacterial mutagen, but knowledge about its metabolism and carcinogenic potential is lacking. Metabolism studies on MeI<i>g</i>Qx and MeIQx were conducted with human and mouse liver microsomes, and recombinant human P450s. DNA binding studies were also investigated in mice to ascertain the genotoxic potential of MeI<i>g</i>Qx in comparison to MeIQx. Both HAAs underwent comparable rates of <i>N</i>-oxidation to form genotoxic <i>N</i>-hydroxylated metabolites with mouse liver microsomes (0.2–0.3 nmol/min/mg protein). The rate of <i>N</i>-oxidation of MeIQx was 4-fold greater than the rate of <i>N</i>-oxidation of MeI<i>g</i>Qx with human liver microsomes (1.7 vs 0.4 nmol/min/mg protein). The rate of <i>N</i>-oxidation, by recombinant human P450 1A2, was comparable for both substrates (6 pmol/min/pmol P450 1A2). MeI<i>g</i>Qx also underwent <i>N</i>-oxidation by human P450s 1A1 and 1B1 at appreciable rates, whereas MeIQx was poorly metabolized by these P450s. The potential of MeI<i>g</i>Qx and MeIQx to form DNA adducts was assessed in female C57BL/6 mice given [¹⁴C]-MeI<i>g</i>Qx (10 μCi, 9.68 mg/kg body wt) or [¹⁴C]-MeIQx (10 μCi, 2.13 mg/kg body wt). DNA adduct formation in the liver, pancreas, and colorectum was measured by accelerator mass spectrometry at 4, 24, or 48 h post-treatment. Variable levels of adducts were detected in all organs. The adduct levels were similar for both HAAs, when adjusted for dose, and ranged from 1 to 600 adducts per 10⁷ nucleotides per mg/kg dose. Thus, MeI<i>g</i>Qx undergoes metabolic activation and binds to DNA at levels that are comparable to MeIQx. Given the high amounts of MeI<i>g</i>Qx formed in cooked meats, further investigations are warranted to assess the carcinogenic potential of this HAA

DNA Adduct Formation of 4-Aminobiphenyl and Heterocyclic Aromatic Amines in Human Hepatocytes.

DNA adduct formation of the aromatic amine, 4-aminobiphenyl (4-ABP), a known human carcinogen present in tobacco smoke, and the heterocyclic aromatic amines (HAAs), 2-amino-9H-pyrido[2,3-b]indole (AαC), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), and 2-amino-3,8-dimethylmidazo[4,5-f]quinoxaline (MeIQx), potential human carcinogens, which are also present in tobacco smoke or formed during the high-temperature cooking of meats, was investigated in freshly cultured human hepatocytes. The carcinogens (10 μM) were incubated with hepatocytes derived from eight different donors for time periods up to 24 h. The DNA adducts were quantified by liquid chromatography-electrospray ionization mass spectrometry with a linear quadrupole ion trap mass spectrometer. The principal DNA adducts formed for all of the carcinogens were N-(deoxyguanosin-8-yl) (dG-C8) adducts. The levels of adducts ranged from 3.4 to 140 adducts per 10(7) DNA bases. The highest level of adduct formation occurred with AαC, followed by 4-ABP, then by PhIP, MeIQx, and IQ. Human hepatocytes formed dG-C8-HAA-adducts at levels that were up to 100-fold greater than the amounts of adducts produced in rat hepatocytes. In contrast to HAA adducts, the levels of dG-C8-4-ABP adduct formation were similar in human and rat hepatocytes. These DNA binding data demonstrate that the rat, an animal model that is used for carcinogenesis bioassays, significantly underestimates the potential hepatic genotoxicity of HAAs in humans. The high level of DNA adducts formed by AαC, a carcinogen produced in tobacco smoke at levels that are up to 100-fold higher than the amounts of 4-ABP, is noteworthy. The possible causal role of AαC in tobacco-associated cancers warrants investigation

REinforcement learning to improve non-adherence for diabetes treatments by Optimising Response and Customising Engagement (REINFORCE): Study protocol of a pragmatic randomised trial

Introduction Achieving optimal diabetes control requires several daily self-management behaviours, especially adherence to medication. Evidence supports the use of text messages to support adherence, but there remains much opportunity to improve their effectiveness. One key limitation is that message content has been generic. By contrast, reinforcement learning is a machine learning method that can be used to identify individuals\u27 patterns of responsiveness by observing their response to cues and then optimising them accordingly. Despite its demonstrated benefits outside of healthcare, its application to tailoring communication for patients has received limited attention. The objective of this trial is to test the impact of a reinforcement learning-based text messaging programme on adherence to medication for patients with type 2 diabetes. Methods and analysis In the REinforcement learning to Improve Non-adherence For diabetes treatments by Optimising Response and Customising Engagement (REINFORCE) trial, we are randomising 60 patients with suboptimal diabetes control treated with oral diabetes medications to receive a reinforcement learning intervention or control. Subjects in both arms will receive electronic pill bottles to use, and those in the intervention arm will receive up to daily text messages. The messages will be individually adapted using a reinforcement learning prediction algorithm based on daily adherence measurements from the pill bottles. The trial\u27s primary outcome is average adherence to medication over the 6-month follow-up period. Secondary outcomes include diabetes control, measured by glycated haemoglobin A1c, and self-reported adherence. In sum, the REINFORCE trial will evaluate the effect of personalising the framing of text messages for patients to support medication adherence and provide insight into how this could be adapted at scale to improve other self-management interventions. Ethics and dissemination This study was approved by the Mass General Brigham Institutional Review Board (IRB) (USA). Findings will be disseminated through peer-reviewed journals, clinicaltrials.gov reporting and conferences. Trial registration number Clinicaltrials.gov (NCT04473326)

Rumination-Focused Cognitive Behavioral Therapy Reduces Rumination and Targeted Cross-network Connectivity in Youth With a History of Depression: Replication in a Preregistered Randomized Clinical Trial

Background: Rumination-focused cognitive behavioral therapy (RF-CBT) is designed to reduce depressive rumination or the habitual tendency to dwell on experiences in a repetitive, negative, passive, and global manner. RF-CBT uses functional analysis, experiential exercises, and repeated practice to identify and change the ruminative habit. This preregistered randomized clinical trial (NCT03859297, R61) is a preregistered replication of initial work. We hypothesized a concurrent reduction of both self-reported rumination and cross-network connectivity between the left posterior cingulate cortex and right inferior frontal and inferior temporal gyri. Methods: Seventy-six youths with a history of depression and elevated rumination were randomized to 10 to 14 sessions of RF-CBT (n = 39; 34 completers) or treatment as usual (n = 37; 28 completers). Intent-to-treat analyses assessed pre-post change in rumination response scale and in functional connectivity assessed using two 5 minute, 12 second runs of resting-state functional magnetic resonance imaging. Results: We replicated previous findings: a significant reduction in rumination response scale and a reduction in left posterior cingulate cortex to right inferior frontal gyrus/inferior temporal gyrus connectivity in participants who received RF-CBT compared with those who received treatment as usual. Reductions were large (z change = 0.84; 0.73, respectively [ps < .05]). Conclusions: This adolescent clinical trial further demonstrates that depressive rumination is a brain-based mechanism that is modifiable via RF-CBT. Here, we replicated that RF-CBT reduces cross-network connectivity, a possible mechanism by which rumination becomes less frequent, intense, and automatic. This National Institute of Mental Health-funded fast-fail study continues to the R33 phase during which treatment-specific effects of RF-CBT will be compared with relaxation therapy