Cancer-Related Somatic Mutations in Transmembrane Helices Alter Adenosine A1 Receptor Pharmacology.

Overexpression of the adenosine A1 receptor (A1AR) has been detected in various cancer cell lines. However, the role of A1AR in tumor development is still unclear. Thirteen A1AR mutations were identified in the Cancer Genome Atlas from cancer patient samples. We have investigated the pharmacology of the mutations located at the 7-transmembrane domain using a yeast system. Concentration-growth curves were obtained with the full agonist CPA and compared to the wild type hA1AR. H78L3.23 and S246T6.47 showed increased constitutive activity, while only the constitutive activity of S246T6.47 could be reduced to wild type levels by the inverse agonist DPCPX. Decreased constitutive activity was observed on five mutant receptors, among which A52V2.47 and W188C5.46 showed a diminished potency for CPA. Lastly, a complete loss of activation was observed in five mutant receptors. A selection of mutations was also investigated in a mammalian system, showing comparable effects on receptor activation as in the yeast system, except for residues pointing toward the membrane. Taken together, this study will enrich the view of the receptor structure and function of A1AR, enlightening the consequences of these mutations in cancer. Ultimately, this may provide an opportunity for precision medicine for cancer patients with pathological phenotypes involving these mutations.

Cancer-related somatic mutations alter adenosine A1 receptor pharmacology-A focus on mutations in the loops and C-terminus.

G protein-coupled receptors (GPCRs) are known to be involved in tumor progression and metastasis. The adenosine A1 receptor (A1 AR) has been detected to be over-expressed in various cancer cell lines. However, the role of A1 AR in tumor development is not yet well characterized. A series of A1 AR mutations were identified in the Cancer Genome Atlas from cancer patient samples. In this study, we have investigated the pharmacology of mutations located outside of the 7-transmembrane domain by using a "single-GPCR-one-G protein" yeast system. Concentration-growth curves were obtained with the full agonist CPA for 12 mutant receptors and compared to the wild-type hA1 AR. Most mutations located at the extracellular loops (EL) reduced the levels of constitutive activity of the receptor and agonist potency. For mutants at the intracellular loops (ILs) of the receptor, an increased constitutive activity was found for mutant receptor L211R5.69 , while a decreased constitutive activity and agonist response were found for mutant receptor L113F34.51 . Lastly, mutations identified on the C-terminus did not significantly influence the pharmacological function of the receptor. A selection of mutations was also investigated in a mammalian system. Overall, similar effects on receptor activation compared to the yeast system were found with mutations located at the EL, but some contradictory effects were observed for mutations located at the IL. Taken together, this study will enrich the insight of A1 AR structure and function, enlightening the consequences of these mutations in cancer. Ultimately, this may provide potential precision medicine in cancer treatment.

Evaluation of Latent Fingerprints for Drug Screening in a Social Care Setting.

Sweat deposited via latent fingerprints (LFPs) was previously used to detect cocaine, opioids, cannabis and amphetamine via a point-of-care test (POCT). This screening method combined non-invasive sampling with a rapid result turnaround to produce a qualitative result outside of the laboratory. We report the novel application of a LFP drug screening test in a social care setting. Clients were tested on either an ad hoc or a routine basis using the POCT DOA114 (Intelligent Fingerprinting Ltd) drug screening cartridge. Screening cutoff values were 45, 35 and 95 pg/fingerprint for benzoylecgonine (BZE), morphine and amphetamine analytes, respectively. Confirmation LFP samples (DOA150, Intelligent Fingerprinting Ltd) and oral fluid (OF) were analyzed using ultra-performance liquid chromatography with tandem mass spectrometry. Thirty-six clients aged 36 ± 11 years participated (53% females). Individuals self-reported alcohol consumption (39%) and smoking (60%). Of 131 screening tests collected over 8 weeks, 14% tested positive for cocaine, 2% tested positive for opioids and 1% tested positive for amphetamine. Polydrug use was indicated in 10% of tests. Of 32 LFP confirmation tests, 63% were positive for cocaine and BZE. Opioids were also detected (31%), with the metabolite 6-monoacetylmorphine (6-MAM) being the most common (16%). In OF, cocaine was the dominant analyte (9%) followed by 6-MAM (5%). On comparing positive LFP screening tests with positive OF samples, we found that 39% and 38% were cocaine and opiate positive, respectively. Of the drugs screened for via the LFP POCT, cocaine was the most prevalent analyte in LFP and OF confirmation samples. The study is a step change in the routine drug screening procedures in a social care setting, especially useful for on-site cocaine detection in clients whose drug use was being monitored. Additionally, testing was easily accepted by clients and social care workers.

Radiofrequency therapy improves exercise capacity of mice with emphysema.

Emphysema is a common phenotype of chronic obstructive pulmonary disease (COPD). Although resection of emphysematous tissue can improve lung mechanics, it is invasive and fraught with adverse effects. Meanwhile, radiofrequency (RF) treatment is an extracorporeal method that leads to tissue destruction and remodeling, resulting in "volume reduction" and overall improvement in lung compliance of emphysematous lungs. Whether these changes lead to improved exercise tolerance is unknown. Here, we investigated the effectiveness of RF treatment to improve the exercise capacity of mice with emphysema. Fifty-two mice (7 weeks of age) were used in this experiment. A bilateral emphysema model was created by intratracheally instilling porcine pancreatic elastase (PPE) (1.5U/100 g body weight). RF treatment (0.5 W/ g body weight) was administered extracorporeally 14 days later and mice were sacrificed after another 21 days. The exercise capacity of mice was measured using a treadmill. Treadmill runs were performed just before PPE instillation (baseline), before RF treatment and before sacrifice. Following sacrifice, lung compliance and mean linear intercept (Lm) were measured and fibrosis was assessed using a modified Ashcroft score. There were 3 experimental groups: controls (instilled with saline, n = 12), emphysema (instilled with porcine pancreatic elastase, PPE, n = 11) and emphysema + treatment (instilled with PPE and given RF, n = 9). At endpoint, the maximum velocity of the emphysema + treatment group was significantly higher than that of the emphysema group, indicating improved exercise tolerance (86.29% of baseline vs 61.69% of baseline, p = 0.01). Histological analysis revealed a significant reduction in emphysema as denoted by Lm between the two groups (median 29.60 µm vs 35.68 µm, p = 0.03). The emphysema + treatment group also demonstrated a higher prevalence of lung fibrosis (≧Grade 3) compared with the emphysema group (11.7% vs 5.4%, p < 0.01). No severe adverse events from RF were observed. RF treatment improved the exercise capacity of mice with emphysema. These data highlight the therapeutic potential of RF treatment in improving the functional status of patients with COPD.

Antenatal smoking and substance-misuse, infant and newborn response to hypoxia.

OBJECTIVES: To determine at the peak age for sudden infant death syndrome (SIDS) the ventilatory response to hypoxia of infants whose mothers substance misused in pregnancy (SM infants), or smoked during pregnancy (S mothers) and controls whose mothers neither substance misused or smoked. In addition, we compared the ventilatory response to hypoxia during the neonatal period and peak age of SIDS. WORKING HYPOTHESIS: Infants of S or SM mothers compared to control infants would have a poorer ventilatory response to hypoxia at the peak age of SIDS. STUDY DESIGN: Prospective, observational study. PATIENT-SUBJECT SELECTION: Twelve S; 12 SM and 11 control infants were assessed at 6-12 weeks of age and in the neonatal period. METHODOLOGY: Changes in minute volume, oxygen saturation, heart rate, and end tidal carbon dioxide levels on switching from breathing room air to 15% oxygen were assessed. Maternal and infant urine samples were tested for cotinine, cannabinoids, opiates, amphetamines, methadone, cocaine, and benzodiazepines. RESULTS: The S and SM infants had a greater decline in minute volume (P = 0.037, P = 0.016, respectively) and oxygen saturation (P = 0.031) compared to controls. In all groups, the magnitude of decline in minute volume in response to hypoxia was higher in the neonatal period compared to at 6-12 weeks (P < 0.001). CONCLUSIONS: Both maternal substance misuse and smoking were associated with an impaired response to a hypoxic challenge at the peak age for SIDS. The hypoxic ventilatory decline was more marked in the neonatal period compared to the peak age for SIDS indicating a maturational effect. Pediatr Pulmonol. 2017;52:650-655. © 2016 Wiley Periodicals, Inc.

Ventilatory Responses to Hypercarbia in Infants of Mothers Who Smoke and Misuse Substances.

The ventilatory response of infants of mothers who smoke and misuse substances and controls to carbon dioxide was assessed at 6-12 weeks and the perinatal period. Infants of mothers who smoke and misuse substances had a dampened response at the peak age of sudden infant death syndrome, greater than in the perinatal period.

Antenatal substance misuse and smoking and newborn hypoxic challenge response.

OBJECTIVES: Infants of smoking (S) and substance misusing (SM) mothers have an increased risk of sudden infant death syndrome. The aim of this study was to test the hypothesis that infants of SM or S mothers compared with infants of non-SM, non-smoking mothers (controls) would have a poorer ventilatory response to hypoxia, which was particularly marked in the SM infants. DESIGN: Physiological study. SETTING: Tertiary perinatal centre. PATIENTS: 21 SM; 21 S and 19 control infants. Infants were assessed before maternity/neonatal unit discharge. INTERVENTIONS: Maternal and infant urine samples were tested for cotinine, cannabinoids, opiates, amphetamines, methadone, cocaine and benzodiazepines. MAIN OUTCOME MEASURES: During quiet sleep, the infants were switched from breathing room air to 15% oxygen and changes in minute volume were assessed. RESULTS: The SM infants had a greater mean increase (p=0.028, p=0.034, respectively) and a greater magnitude of decline (p<0.001, p=0.018, respectively) in minute volume than the S infants and the controls. The rate of decline in minute volume was greater in the SM infants (p=0.008) and the S infants (p=0.011) compared with the controls. CONCLUSIONS: Antenatal substance misuse and smoking affect the infant's ventilatory response to a hypoxic challenge.

Pregnancy outcomes in substance-misusing pregnant women: A 10-year retrospective study.

In this ten-year retrospective study we were able to show that Maltese substance-misusing mothers (SMMs) have significantly different socio-biological characteristics from mothers in the general population which seem to impact on neonatal outcomes. Significant differences from the general population were noted in the SMM's age, age at first pregnancy, marital status, educational background, number of cigarettes smoked, number of offspring and weight gained in pregnancy. SMMs booked their pregnancy in hospital later than the general population. On the other hand their infants had a lower mean birth weight and head circumference, had lower Apgar scores, were of lesser gestational age and premature. They were less breast-fed and stayed longer in hospital after their birth.

Ventilatory response to hypercarbia in newborns of smoking and substance-misusing mothers.

RATIONALE: Infants of mothers who smoked (S) or substance misused (SM) during pregnancy have an increased risk of sudden infant death syndrome (SIDS). OBJECTIVES: To test the hypothesis that infants of S and SM mothers compared with infants of non-substance-misusing, nonsmoking mothers (control subjects) would have a reduced ventilatory response to hypercarbia and that any reduction would be greater in the SM infants. METHODS: Infants were assessed before maternity/neonatal unit discharge. Maternal and infant urine samples were obtained and tested for cotinine, cannabinoids, opiates, amphetamines, methadone, cocaine, and benzodiazepines. MEASUREMENTS AND MAIN RESULTS: Respiratory flow and Vt were measured using a pneumotachograph inserted into a face mask placed over the infant's mouth and nose. The ventilatory responses to three levels of inspired carbon dioxide (0 [baseline], 2, and 4% CO2) were assessed. Twenty-three SM, 34 S, and 22 control infants were assessed. The birth weight of the control subjects was higher than the SM and S infants (P = 0.017). At baseline, SM infants had a higher respiratory rate (P = 0.003) and minute volume (P = 0.007) compared with control subjects and S infants. Both the SM and S infants had a lower ventilatory response to 2% (P < 0.001) and 4% (P < 0.001) CO2 than the control subjects. The ventilatory response to CO2 was lower in the SM infants compared with the S infants (P = 0.009). CONCLUSIONS: These results are consistent with infants of smoking mothers and substance misuse/smoking mothers having a dampened ventilatory response to hypercarbia, which is particularly marked in the latter group.

Prevalence of the tobacco product dokha among high school students in Dubai.

BACKGROUND: Dokha is a mixed tobacco product, smoked through a pipe, traditional in Arabic culture and popular in the UAE. User forums suggest this product may contain higher levels of nicotine and potentially psychoactive properties. However, there have been few published studies on dokha prevalence and health effects. The present study provides initial data on prevalence of use among high school students resident in Dubai, UAE. METHODS: 416 students were recruited from five English curriculum schools. Respondents completed a questionnaire measuring current and lifetime use of tobacco in the form of cigarettes, water-pipes and dokha. Current tobacco users completed the 'Hooked on Nicotine Checklist for signs of dependency. RESULTS: Data from 394 subjects were analyzed with mean age 16.9 years. Regular use (weekly or daily) of tobacco had relatively high prevalence (23.4%) compared to international data and was driven by use of the local tobacco, dokha. CONCLUSION: Dokha forms a commonly used tobacco product among young people in the UAE from both Arab and Western national groups. Little is currently known about the health and dependence risks of this product. The paper adds to the few studies calling for timely research into this and other emerging tobacco products.

Cannabis use: a perspective in relation to the proposed UK drug-driving legislation.

With regard to THC (Δ(9)-tetrahydrocannabinol), the main psychoactive constituent identified in the plant Cannabis sativa L, several facts are indisputable. Cannabis remains the most commonly used drug in the UK among those who reported driving under the influence of illegal drugs in the previous 12 months. There is a significant dose-related decrement in driving performance following cannabis use; raised blood THC concentrations are significantly associated with increased traffic crash and death risk. When cannabis and alcohol are detected together, there is a greater risk to road safety than when either drug is used alone. Patterns of use are important when interpreting blood concentration data: Smoking infrequently a single cannabis cigarette leads to peak plasma THC concentrations (21-267 µg/L) causing acute intoxication. In habitual, daily users, plasma THC concentrations range from 1.0 to 11.0 µg/L and are maintained by sequestration of the drug from the tissues. These facts undoubtedly make setting thresholds for drug-driving legislation difficult but there is clearly a case for cannabis. Determining minimum blood THC concentrations at which a driver becomes sufficiently impaired to be unable to safely drive a vehicle is of particular concern given the increasing medicinal use of the drug. Internationally legislation for driving under the influence of drugs (DUID) is based on either a proof of impairment or a per se approach. For the latter this can be either zero-tolerance or based on concentration limits such as those used for alcohol. The different approaches are considered against current scientific evidence.

A comparison of smoking behaviour characteristics between Caucasian smokers in the United Kingdom and Malay smokers in Malaysia.

OBJECTIVES: There is evidence that smoking behaviour differs by ethnicity. This study aims to compare smoking behaviour characteristics between Caucasian and Malay smokers. METHODS: A cross sectional survey, involving 175 smokers attending smoking cessation clinics at the Institute of Psychiatry, London, United Kingdom and University Malaya, Kuala Lumpur, Malaysia between May 2005 and February 2007. Data on demographics, smoking history, nicotine dependence and smoking behaviour were collected. RESULTS: All participants were males, mean age 30.7 ± 10.3 years. Caucasians initiated smoking significantly earlier (mean age 14.8 ± 2.8 years) (p = 0.001) and smoked regularly significantly earlier (mean age 17.3 ± 3.5) (p = 0.003) than Malays (mean starting age 16.9 ± 4.4 years and mean age regular use 19.5 ± 4.5 years), respectively. Caucasians smoked less for social integration than Malays (p = 0.03) but smoked more for regulation of negative affect than Malays (p = 0.008) and smoked more for hedonism than Malays (p < 0.001). CONCLUSION: Malays smoke as a means of socially integrating. This has important public health implications. Social reasons and the social environment play a role in smoking uptake, smoking maintenance and smoking cessation and this should be borne in mind for strategies planning to promote smoking cessation.

Follow-up of provision of inpatient treatment for UK Healthcare Professionals with alcohol dependence: snapshot of a pilot specialist National Health Service.

Alcohol dependent healthcare professionals (AHCPs) entering a specialized National Health Service (NHS) inpatient treatment facility in southeast London, UK, were investigated. A retrospective case-note analysis was conducted, supplemented with a postal questionnaire. Twenty-three mature (mean age 46.7 years; 13 male) AHCPs, 82% of admissions, with a mean duration of alcohol dependence of 16.5 years were followed-up at an average of 26 months postdischarge. Half of the sample met criteria for lifetime psychiatric disorder. Physical illness was also a prominent problem. At follow-up, six professionals reported continuous abstinence since discharge. For those who continued drinking, there was a significant reduction in daily quantity consumed. This study suggests that the absence of an adequate treatment infrastructure and administrative resources have a major impact on access to treatment services and outcomes for addicted healthcare professionals. There is a need for supported and dedicated services for this group within the UK National Health Service. The study's limitations are noted.

Persistence of heroin use despite methadone treatment: poor coping self-efficacy predicts continued heroin use.

AIM: To evaluate the association between coping self-efficacy and persistent use of heroin by patients enrolled in a methadone treatment program. DESIGN AND METHODS: Cross-sectional survey. One hundred and ninety-one patients attending outpatient methadone clinics in South-East England, United Kingdom. Validated questionnaires were used to assess drug use (Maudsley Addiction Profile), alcohol use (Alcohol Use Disorders Identification Test), mental health (Hospital Anxiety and Depression Scale) and coping self-efficacy (brief 8-item Drug Taking Confidence Questionnaire). RESULTS: Half of the participants (95/191) reported heroin use in the preceding 14-day period. Heroin use during methadone treatment was associated with financial problems (P = 0.008), spending time with other drug users (P < 0.001), cocaine use (P = 0.002), low mood (P = 0.002) and dissatisfaction with the daily methadone dose (P = 0.014). Compared with 'Heroin-abstinent' patients, the 'Heroin' group reported significantly lower mean coping self-efficacy scores (t = 9.8, d.f. = 182, P < 0.001, effect size 1.17). After correcting for the effects of co-variants in a logistic regression model, the main determinants of persistent heroin use were 'coping self-efficacy' [B -0.05; standard error (SE) 0.008; Wald 36.6; odds ratio (OR) 0.95, 95% confidence interval (CI) 0.94, 0.97; P < 0.001] and 'dissatisfaction with methadone dose' (B 0.93; SE 0.46; Wald 4.1; OR 2.5, 95% CI 1.03, 6.25; P = 0.042). Satisfaction with methadone dose showed no association with self-efficacy. DISCUSSION AND CONCLUSIONS; While heroin use during methadone treatment can partly be explained by inadequate dosing, our data suggest a more complex picture with significant contribution from poor coping self-efficacy. Efforts aimed at enhancing and maintaining the patients' self-efficacy and social skills are likely to improve heroin and other drug use outcomes with added benefits for treatment completion rates and the throughput of methadone programs.

Changes to methadone clearance during pregnancy.

OBJECTIVE: Measurement of plasma methadone concentration to investigate the rate of clearance of methadone prescribed for heroin dependence in the first, second and third trimesters of pregnancy. A secondary objective was to evaluate the outcome of pregnancy. METHODS: Longitudinal within subject study of nine pregnant opioid dependent subjects prescribed methadone at the Leeds Addiction Unit, an outpatient community based treatment centre. Plasma concentration versus time data for methadone was collected during each trimester and post-partum for our subjects. Data was available for the first and second trimesters for 4/9 cases. All but one of the subjects provided data during the third trimester and data post-partum was collected from three respondents. Measurements of methadone levels in plasma were carried out using high performance liquid chromatography (HPLC). RESULTS: Trough mean plasma methadone concentrations reduced as the pregnancies progressed from 0.12 mg/L (first trimester) to 0.07 mg/L (third trimester). The weight-adjusted clearance rates gradually increased from a mean of 0.17 to 0.21 L/hr/kg during pregnancy, although patterns differed substantially between the nine women. An assessment of relative clearance of methadone using two patients for whom we have had all three CL values (trimester 1-3) demonstrated notable change of CL (P = 0.056) over time. Eight of our subjects delivered (3 males), within two weeks of their due date the ninth (male) was premature (21 days). The mean length of gestation was 39.7 weeks (SD = 10 days) and none of the neonates met criterion for 'low birth weight' mean = 3094, SD = 368 g). Five neonates spent time (0.5-28 days) in a special care baby unit (SCUBU) and 4 of these displayed signs of methadone withdrawal. CONCLUSIONS: General Practitioners and hospital doctors should recognise the significant benefits of prescribing methadone for heroin-dependent women during pregnancy. We recommend that if a pregnant opioid user complains of methadone withdrawal symptoms (i.e. that the methadone dose does not "hold" them) the prescribing clinician takes this observation seriously and considers a more detailed assessment. Further work on key factors undergoing changes during pregnancy accounting for differences in methadone metabolism in the mother, fetus and neonate are required.

The effect of tobacco smoking on subjective symptoms of inadequacy ("not holding") of methadone dose among opiate addicts in methadone maintenance treatment.

The objective of this study was to examine the relationship between subjective symptoms of inadequacy of methadone dose (not feeling "held") and tobacco smoking in patients in methadone maintenance treatment (MMT). This was a cross-sectional study of smoking behaviour, investigating subjective, physiological and psychological symptoms. The study took place in a community-based methadone maintenance clinic of a psychiatric teaching hospital in South London. Fifty adult opiate addicts (37 males and 13 females) were on a stable daily methadone dose; the number of cigarettes smoked during the day and previous day of investigation, salivary cotinine measurements and carbon monoxide (CO) from expired air were measured. The Methadone Symptom Checklist (MSC) was used to score withdrawal symptoms encountered in patients not feeling "held" during MMT The Hamilton Anxiety Score was also used. The prevalence of tobacco-smoking was high (98%), with two-thirds (68%) smoking self-fabricated cigarettes ("roll-ups"). Scores from rating scales measuring symptoms of not being "held" correlated with number of cigarettes smoked the previous day (p < 0.05). A similar correlation was found with the Hamilton Anxiety Score. However, there was no correlation between rating scale scores and either salivary cotinine concentration or CO from expired air. Methadone patients who smoke more are significantly more likely to report problems of not feeling "held" by their methadone dose and they also show a higher level of anxiety. However, this increased cigarette consumption is not reflected in increased salivary continine levels or levels of CO in expired air, and it may be that the raised level of anxiety leads to a smoking-pattern consisting of frequent lighting-up of cigarettes or "roll-ups" which are consumed incompletely and/or not smoked by inhalation.