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Refereed/Peer-reviewed We aimed to develop population pharmacokinetic/pharmacodynamic (PK/PD) models that can effectively describe ketamine and norketamine PK/PD relationships for Montgomery–Åsberg Depression Rating Scale (MADRS) scores, blood pressure (BP), and heart rate (HR) following i.v., s.c., and i.m. ketamine administration in patients with treatment-refractory depression. Ketamine PK/PD data were collected from 21 treatment-refractory depressed participants who received ketamine (dose titration 0.1–0.5 mg/kg as single doses) by i.v., s.c., or i.m. administration. Model development used nonlinear mixed effect modeling. Ketamine and norketamine PK were best described using two-compartment models with first-order absorption after s.c. and i.m. administration. Estimated ketamine bioavailability after i.m. and s.c. was ~ 64% with indistinguishable first-order absorption rate constants. Allometric scaling of body weight on all clearance and volumes of distribution improved the model fit. The delay in the concentration-response relationship for MADRS scores was best described using a turnover model (turnover time ~ 42 hours), whereas for the BP and HR rates this was an immediate effect model. For all PD effects, ketamine alone was superior to models with norketamine concentration linked to an effect. No covariates were identified for PD effects. The estimated half-maximal effective concentration from the MADRS score, BP, and HR were 0.44, 468, and 7,580 ng/mL, respectively. The integrated population models were able to effectively describe the PK/PD relationships for MADRS scores, BP, and HR after i.v., s.c., and i.m. ketamine administration. These findings allow for a deeper understanding of the complex relationships between route of ketamine administration and clinical response and safety.

Objective Pain severity and opioid requirements in the postoperative period show substantial and clinically significant inter-patient variation due mainly to factors such as age, surgery type, and duration. Genetic factors have not been adequately assessed except for the neuronal OPRM1 rs1799971 and COMT rs4680, whereas the contribution of innate immune signaling pathway genetics has seldom been investigated. Setting Hospital surgical ward. Subjects Women (107 Indian, 184 Malay, and 750 Han Chinese) undergoing total hysterectomy surgery. Methods Morphine consumption, preoperative pain, and postoperative pain were evaluated in relation to genetic variability comprising 19 single-nucleotide polymorphisms (SNPs) in 14 genes involved in glial activation, inflammatory signaling, and neuronal regulation, plus OPRM1 (1 SNP) and COMT (3 SNPs). Results Pre- and postoperative pain and age were associated with increased and decreased morphine consumption, respectively. In Chinese patients, only 8% of the variability in consumption could be explained by these nongenetic and genetic (BDNF, IL1B, IL6R, CRP, OPRM1, COMT, MYD88) factors. However, in Indian patients, 41% of morphine consumption variability could be explained by age (explaining Conclusions This is the highest known value reported for genetic contributions (38%) to morphine use in the acute postoperative pain setting. Our findings highlight the need to incorporate both genetic and nongenetic factors and consider ethnicity-dependent and nonadditive genotypic models in the assessment of factors that contribute to variability in opioid use.

Opioids remain the major drug class for the treatment of acute, chronic and cancer pain, but have major harmful effects such as dependence and opioid-induced ventilatory impairment. Although no new typical opioids have come onto the market in the past almost 50 years, a plethora of new innovative formulations has been developed to meet the clinical need. This review is intended to shed light on new understanding of the molecular pharmacology of opioids, which has arisen largely due to the genomic revolution, and what new drugs may become available in the coming years. Atypical opioids have and are being developed which not only target the mu opioid receptor but other targets in the pain pathway. Biased mu agonists have been developed but remain ‘unbiased’ clinically. The contribution of drugs targeting non-mu opioid receptors either alone or as heterodimers shows potential promise but remains understudied. That gene splice variants of the mu opioid receptor produce multiple receptor isoforms in different brain regions, and may change with pain chronicity and phenotype, presents new challenges but also opportunities for precision pain medicine. Finally, that opioids also have pro-inflammatory effects not aligned with mu opioid receptor binding affinity implicates a fresh understanding of their role in chronic pain, whether cancer or non-cancer. Hopefully, a new understanding of opioid analgesic drug action may lead to new drug development and better precision medicine in acute and chronic pain relief with less patient harm.

Papua New Guinea (PNG) has a high HIV/AIDS prevalence and very high frequency of the CYP2B6 c.516G>T (rs3745274) variant. We have conducted the first investigation of the impact of c.516G>T and patient demographics on plasma efavirenz (EFV) and 8‐hydroxyefavirenz (8OH‐EFV) concentrations, metabolic ratio (8OH‐EFV/EFV) (MR), and their association with adverse effects, in PNG patients with HIV/AIDS. For 156 PNG patients with HIV/AIDS taking EFV 600 mg/day (for 3–156 months), plasma EFV and 8OH‐EFV concentrations were quantified, CYP2B6 c.516G>T genotyped, and demographic and self‐reported adverse effects data recorded. Genotype differences in EFV and 8OH‐EFV concentrations, MR, and percent within therapeutic range (1000–4000 ng/ml) were examined, in addition to EFV and 8OH‐EFV concentration differences between patients experiencing adverse effects. CYP2B6 c.516T allele frequency was 53%. Plasma EFV (p

Accurate quantification of efavirenz metabolites in patient samples is required to investigate their potential contribution to efavirenz adverse events. This study aimed to validate a LC-MS/MS method to quantify and investigate the stability of efavirenz and metabolites in human plasma. Compounds were extracted from plasma by supported liquid extraction and resolved on a C18 column. Validation was performed following FDA bioanalytical method validation guidelines. Stability under common conditions of sample pre-treatment and storage were assessed. Efavirenz and 8-hydroxyefavirenz were stable for all conditions tested. 7-Hydroxyefavirenz and 8,14-dihydroxyefavirenz were not stable in plasma at room temperature for 24 h (46%-69% loss), -20°C for 90 days (17%-50% loss), or 60°C for 1 h (90%-95% loss). Efavirenz and 8-hydroxyefavirenz concentrations in HIV/AIDS patient (n=5) plasma prepared from pre-treated (60°C for 1 h) whole blood varied from 517-8564 ng/mL and 131-813 ng/mL, respectively. 7-Hydroxyefavirenz and 8,14-dihydroxyefavirenz concentrations were below validated lower limits of quantification (0.25 and 0.5 ng/mL, respectively), most likely due to sample pre-treatment. This is the first report of 7-hydroxyefavirenz and 8,14-dihydroxyefavirenz instability under conditions commonly used in preparation of samples from HIV/AIDS patients. Alternative biosafety measures to heat pre-treatment must therefore be used for accurate quantification of plasma 7-hydroxyefavirenz and 8,14-dihydroxyefavirenz.

Aims: Long-term use of the immunosuppressant tacrolimus is limited by nephrotoxicity. Following renal transplantation, the risk of nephrotoxicity may be determined more by allograft than by blood tacrolimus concentrations, and thus may be affected by donor CYP3A5 and ABCB1 genetics. Little is known regarding factors that determine tacrolimus intrarenal exposure. Methods: This study investigated the relationship between trough blood (C0Blood) and allograft (CGraft) tacrolimus concentrations and tacrolimus dose, haematocrit, genetics, acute nephrotoxicity, rejection status, delayed graft function, and time post-transplant. C0Blood and CGraft were quantified in 132 renal transplant recipients together with recipient and donor CYP3A5 (rs776746) and ABCB1 3435 (rs1045642) genotypes. Results: C0Blood ranged from 2.6 to 52.3 ng/mL and CGraft from 33 to 828 pg/mg tissue. Adjusting for dose, recipients who were CYP3A5 expressors had lower C0Blood compared to nonexpressors, whilst delayed graft function was associated with higher C0Blood. Linear regression showed that the significant predictors of CGraft were C0Blood (point-wise P = 7 × 10−10), dose (P = .004) acute nephrotoxicity (P = .002) and an interaction between C0Blood and acute tacrolimus nephrotoxicity (P = .0002), with an adjusted r2 = 0.35 and no contribution from donor or recipient CYP3A5 or ABCB1 genotype. The association between CGraft and acute nephrotoxicity depended on one very high CGraft (828 pg/mg tissue). Conclusions: Recipient and donor CYP3A5 and ABCB1 3435C>T genotypes are not determinants of allograft tacrolimus exposure in kidney transplant recipients. However, tacrolimus dose and C0Blood were significant predictors of CGraft, and the relationship between C0Blood and CGraft appeared to differ in the presence or absence of acute nephrotoxicity. Refereed/Peer-reviewed

Background Persistent postoperative pain (PPP) is common after total knee arthroplasty (TKA). The primary aim of this prospective cohort study was to identify important predictors of moderate to severe PPP 6 and 12 months after TKA. Methods Consenting patients (n=300) undergoing primary unilateral TKA attended a preoperative session to collect clinical information (age, gender, BMI, preoperative knee pain, comorbid pain, likely neuropathic pain) and psychological variables (depression, anxiety, catastrophising, expected pain). Quantitative sensory testing (pressure pain thresholds, temporal summation, conditioned pain modulation) was performed, and blood samples were obtained for subsequent genotyping of OPRM1 and COMT. Acute postoperative pain was measured at rest and during movement. Surgical factors (surgery time, patella resurfacing, anaesthetic type) were collected after operation. Follow-up questionnaires were sent 6 and 12 months after surgery. Multivariate logistic regression was used to identify predictors of PPP. Results The prevalence of moderate to severe PPP was 21% (n=60) and 16% (n=45) 6 and 12 months after surgery, with 55% (n=33) and 60% (n=31) of PPP likely neuropathic in nature. At 6 months, a combination of preoperative pain intensity, expected pain, trait anxiety, and temporal summation (Akaike information criterion, 309.9; area under receiver operating characteristic (ROC) curve, 0.70) was able to correctly classify 66% of patients into moderate to severe PPP and no to mild PPP groups. At 12 months, preoperative pain intensity, expected pain, and trait anxiety (Akaike information criterion, 286.8; area under ROC curve, 0.66) correctly classified 66% of patients. Conclusions Findings from this study highlight several factors that may be targeted in future intervention studies to reduce the development of PPP. Trial registry number ACTRN12612001089820.

Tacrolimus (TAC) is a first-line immunosuppressant used to prevent organ rejection after kidney transplantation. There is large inter-individual variability in its pharmacokinetics. Single nucleotide polymorphisms (SNPs) in genes encoding TAC metabolizing enzymes cytochromes P450 3A4/5 (CYP3A4/5), P-glycoprotein efflux transporter (ABCB1), their expression regulator pregnane X receptor (NR1I2) and CYP3A co-factor cytochrome P450 reductase (POR) have been studied for their effects on tacrolimus disposition. However, except for CYP3A5*3, controversies remain about their roles in predicting dose-adjusted trough blood TAC concentrations (C0 /D). This study aimed to investigate the effects of ABCB1 (61A>G, 1199G>A, 1236C>T, 2677G>T and 3435C>T), CYP3A4*22, CYP3A5*3, NR1I2 (8055C>T, 63396C>T and -25385C>T) and POR*28 SNPs on TAC C0 /D. In total, 165 kidney transplant recipients were included in this study. SNPs were genotyped by probe-based real-time polymerase chain reaction. Associations between log-transformed whole blood TAC C0 /D (measured at 1 and 3 months post-transplant) and genotypes/haplotypes were assessed by linear mixed effects analysis, controlling for age, sex and haematocrit. It was observed that CYP3A5 expressors (*1/*1 + *1/*3) (p = 5.5 × 10-16 ) and ABCB1 61G allele carriers (p = 0.001) had lower log-transformed TAC C0 /D (56% and 26% lower geometric mean TAC C0 /D, respectively) and accounted for approximately 30% and 4%, respectively, of log-transformed TAC C0 /D variability in the first 3 months post-transplant. In conclusion, CYP3A5*3 is a major, and ABCB1 61A>G is a novel, although minor, genetic factor affecting TAC C0 /D in kidney transplant recipients.

There are inconsistent findings regarding the relationship between trough whole blood tacrolimus concentration (TAC C0) and acute kidney rejection in recipients undergoing TAC therapeutic drug monitoring (TDM). However, studies have not always assessed TAC C0 at the time of rejection or accounted for variability in hematocrit. Therefore, this study aimed to investigate the temporal relationship between TAC C0 and acute rejection, including when accounting for variation in hematocrit.For 38 recipients who developed biopsy-proven acute rejection (BPAR) in the first 14 days after kidney transplantation, daily TAC C0 from TDM and hematocrit was collected from case notes. Differences in log10-transformed TAC C0 between the day of BPAR (log Cr), 1 day before BPAR (log Cr-1), and 2 days before BPAR (log Cr-2) and the combined median concentrations for the days preceding these (log Cprior) were examined by repeated-measures analysis of variance with Dunnett post hoc testing. Generalized linear mixed-effects regression (glmer) examined the ability of TAC C0 to predict acute rejection episodes with and without controlling for hematocrit.Log Cr-1 [mean difference (95% confidence interval) = -0.13 (-0.21 to -0.048), post hoc P = 0.002] and log Cr [-0.13 (-0.24 to -0.025), post hoc P = 0.013] were significantly lower than log Cprior. TAC C0 was a significant (P = 0.0078) predictor of rejection episodes (area under the receiver operating characteristic curve = 0.79) only in glmer models accounting for variability in hematocrit.In recipients who developed BPAR, there was a significant temporal relationship between TAC C0 and BPAR incidence under TAC TDM that may not be detected in cross-sectional studies, especially if variability in hematocrit is not addressed. This supports a TAC C0-rejection relationship, which differs between recipients, and may explain why some recipients do or do not experience rejection within or below the TDM range, respectively. However, studies with larger sample sizes are needed to confirm this finding.

Phenytoin drug reaction with eosinophilia and systemic symptoms (DRESS) in 3 Aboriginal Australians positive for HLA‐B*56:02 has been previously reported. We report the allele frequency of HLA‐B*56:02 in 2 South Australian populations, 1 Aboriginal (4.8%, 95% confidence interval 2.4–7.8%) and the other European (0%). We compared the frequency with publicly available information on HLA‐B*56:02 status in other Indigenous Australian (n = 4) and European Australian cohorts (n = 1). In the Indigenous Australian cohorts, HLA‐B*56:02 allele frequency ranged from 1.3 to 19%. We also describe an additional case of phenytoin DRESS (RegiSCAR DRESS score 7) in an Aboriginal Australian that was associated with HLA‐B*56:02 and with CYP2C9*1/*3 genotype. In Aboriginal Australians, phenytoin DRESS appears distinctly linked to HLA‐B*56:02 with an allele carriage rate substantially higher than in Europeans, but also with considerable regional variation. Investigations of human leucocyte antigen and other contributing genes and severe adverse drug reactions in understudied non‐European populations are required to optimize safe medication use and inform risk mitigation strategies.

Although several genetic factors have been associated with postsurgical morphine requirements, those involving the innate immune system and cytokines have not been well investigated. The aim of this study was to investigate the contribution of genetic variability in innate immune signalling pathways to variability in morphine dosage after elective caesarean section under spinal anaesthesia in 133 Indian, 230 Malay, and 598 Han Chinese women previously studied. Twenty single nucleotide polymorphisms in 14 genes involved in glial activation (TLR2, TLR4, MYD88, MD2), inflammatory signalling (IL2, IL6, IL10, IL1B, IL6R, TNFA, TGFB1, CRP, CASP1), and neuronal regulation (BDNF) were newly investigated, in addition to OPRM1, COMT, and ABCB1 genetic variability identified previously. Postsurgical patient-controlled analgesia morphine use (mg/24 hours) was binned into 6 normally distributed groups and scored 0 to 5 to facilitate step-down multiple linear regression analysis of genetic predictors, controlling for ethnicity and nongenetic variables. Ethnicity, OPRM1 rs1799971 (increased), TLR2 rs3804100 (decreased), and an interaction between ethnicity and IL1B rs1143634 (increased), predicted 9.8% of variability in morphine use scores in the entire cohort. In the Indian cohort, 14.5% of the variance in morphine use score was explained by IL1B rs1143634 (increased) and TGFB1 rs1800469 (decreased). In Chinese patients, the incidence of postsurgical pain was significantly higher in variant COMT rs4680 genotypes (P = 0.0007) but not in the Malay or Indian cohorts. Innate immune genetics may contribute to variability in postsurgical opioid requirements in an ethnicity-dependent manner.

Background CYP3A4/5 and P-glycoprotein (P-gp, ABCB1) affect tacrolimus (TAC) exposure in T cells and kidney cells. Genetic variability of these genes has been widely studied for effects on acute rejection and kidney function after transplantation, but findings remain contradictory. In addition, cytochrome P450 reductase (POR) is important for CYP3A4/5 activity, and the pregnane X receptor (NR1I2) regulates CYP3A4/5 and P-gp expression. However, the relationship between POR and NR1I2 genetics and acute rejection and kidney function has not been extensively investigated. Objective The aim of this study was to investigate the effect of ABCB1 (61A>G, 1199G>A, 1236C>T, 2677G>T, 3435C>T), CYP3A4*22, CYP3A5*3, NR1I2 (8055C>T, 63396C>T) and POR*28 genotypes/haplotypes on acute rejection and kidney function in the first 3 months after transplant. Participants and methods The study included 165 kidney transplant recipients, who received TAC, mycophenolate and prednisolone, and 129 donors. TAC dose was adjusted to target trough blood concentrations of 8-15 ng/ml by therapeutic drug monitoring. Recipient and donor genotype/haplotype differences in acute rejection incidence within the first 2 weeks after transplant were assessed by logistic regression, adjusting for induction therapy, human leucocyte antigen mismatches, kidney transplant number, peak panel-reactive antibodies and donor type. Recipient and donor genotype/haplotype differences in estimated glomerular filtration rate in the first 3 months after transplant were assessed by linear mixed effects analysis, adjusting for acute rejection, delayed graft function and donor type. Results No genetic factors significantly affected acute rejection or estimated glomerular filtration rate after correction for multiple comparisons (P>0.004). Conclusion Recipient and donor dispositional genetics had no significant effect on short-term clinical outcomes in kidney transplant patients receiving TAC therapeutic drug monitoring.

Papua New Guinea (PNG) can be roughly divided into highland, coastal and island peoples with significant mitochondrial DNA differentiation reflecting early and recent distinct migrations from Africa and East Asia, respectively. Infectious diseases such as tuberculosis, malaria and HIV severely impact on the health of its peoples for which drug therapy is the major treatment and pharmacogenetics has clinical relevance for many of these drugs. Although there is generally little information about known single nucleotide polymorphisms in the population, in some instances, their frequencies have been shown to be higher than anywhere worldwide. For example, CYP2B6*6 is over 50%, and CYP2C19*2 and *3 are over 40 and 25%, respectively. Conversely, CYP2A6*9, 2B6*2, *3, *4 and *18, and 2C8*3 appear to be much lower than in Whites. CYP2D6 known variants are unclear, and for phase II enzymes, only UGT2B7 and UGT1A9 data are available, with variant frequencies either slightly lower than or similar to Whites. Although almost all PNG people tested are rapid acetylators, but which variant(s) define this phenotype is not known. For HLA-B*13:01, HLA-B*35:05 and HLA-C*04:01, the frequencies show some regioselectivity, but the clinical implications with respect to adverse drug reactions are not known. There are minimal phenotype data for the CYPs and nothing is known about drug transporter or receptor genetics. Determination of genetic variants that are rare in Whites or Asians but common in PNG people is a topic of both scientific and clinical importance, and further research needs to be carried out. Optimizing the safety and efficacy of infectious disease drug therapy through pharmacogenetic studies that have translation potential is a priority.

1. Imatinib is metabolized to N-desmethyl imatinib by CYPs 3A4 and 2C8. The effect of CYP2C8*3 genotype on N-desmethyl imatinib formation was unknown. 2. We examined imatinib N-demethylation in human liver microsomes (HLMs) genotyped for CYP2C8*3, in CYP2C8*3/*3 pooled HLMs and in recombinant CYP2C8 and CYP3A4 enzymes. Effects of CYP-selective inhibitors on N-demethylation were also determined. 3. A single-enzyme Michaelis-Menten model with autoinhibition best fitted CYP2C8*1/*1 HLM (n = 5) and recombinant CYP2C8 kinetic data (median ± SD Ki = 139 ± 61 µM and 149 µM, respectively). Recombinant CYP3A4 showed two-site enzyme kinetics with no autoinhibition. Three of four CYP2C8*1/*3 HLMs showed single-enzyme kinetics with no autoinhibition. Binding affinity was higher in CYP2C8*1/*3 than CYP2C8*1/*1 HLM (median ± SD Km = 6 ± 2 versus 11 ± 2 µM, P=0.04). CYP2C8*3/*3 (pooled HLM) also showed high binding affinity (Km = 4 µM) and single-enzyme weak autoinhibition (Ki = 449 µM) kinetics. CYP2C8 inhibitors reduced HLM N-demethylation by 47-75%, compared to 0-30% for CYP3A4 inhibitors. 4. In conclusion, CYP2C8*3 is a gain-of-function polymorphism for imatinib N-demethylation, which appears to be mainly mediated by CYP2C8 and not CYP3A4 in vitro in HLM.

The aims of this study were to determine the effects of the CYP2C8*3 and *4 polymorphisms on imatinib metabolism and plasma imatinib concentrations in chronic myeloid leukaemia (CML) patients.We genotyped 210 CML patients from the TIDELII trial receiving imatinib 400-800 mg/day for CYP2C8*3 (rs11572080, rs10509681) and *4 (rs1058930). Steady-state trough total plasma N-desmethyl imatinib (major metabolite):imatinib concentration ratios (metabolic ratios) and trough total plasma imatinib concentrations were compared between genotypes (one-way ANOVA with Tukey post hoc).CYP2C8*3 (n = 34) and *4 (n = 15) carriers had significantly higher (P 0.01) and lower (P 0.01) metabolic ratios, respectively, than CYP2C8*1/*1 (n = 147) patients (median ± standard deviation: 0.28 ± 0.08, 0.18 ± 0.06 and 0.22 ± 0.08, respectively). Plasma imatinib concentrations were consequently 50% higher for CYP2C8*1/*4 than for CYP2C8*1/*1 and CYP2C8*3 carriers (2.18 ± 0.66 vs. 1.45 ± 0.74 [P 0.05] and 1.36 ± 0.98 μg/mL [P 0.05], respectively).CYP2C8 genotype significantly alters imatinib metabolism in patients through gain- and loss-of-function mechanisms.

The electroencephalogram (EEG) records the electrical activity of the brain and enables effects of anaesthetic drugs on brain functioning to be monitored. Identification of genes contributing to EEG variability during anaesthesia is important to the clinical application of anaesthesia monitoring and may provide an avenue to identify molecular mechanisms underlying the generation and regulation of brain oscillations. Central immune signalling can impact neuronal activity in the brain and accumulating evidence suggests an important role for cytokines as neuronal modulators. We tested 21 single-nucleotide polymorphisms (SNPs) in immune-related genes for associations with three anaesthesia-induced EEG patterns; spindle amplitude, delta power and alpha power, during general anaesthesia with desflurane in 111 patients undergoing general, gynaecological or orthopaedic surgery. Wide inter-patient variability was observed for all EEG variables. MYD88 rs6853 (p = 6.7 × 10(-4)) and IL-1β rs1143627 in conjunction with rs6853 (p = 1.5 × 10(-3)) were associated with spindle amplitude, and IL-10 rs1800896 was associated with delta power (p = 1.3 × 10(-2)) suggesting involvement of cytokine signalling in modulation of EEG patterns during desflurane anaesthesia. BDNF rs6265 was associated with alpha power (p = 3.9 × 10(-3)), suggesting differences in neuronal plasticity might also influence EEG patterns during desflurane anaesthesia. This is the first study we are aware of that has investigated genetic polymorphisms that may influence the EEG during general anaesthesia.

Anti-addiction agents are important psychopharmacological agents used to treat addiction to opioids (illicit and licit), alcohol and nicotine. The goal of treatment is to eliminate further use of the addicted drug by either preventing addicted drug reward or managing withdrawal symptoms experienced when the addicted drug is no longer used. Treatment success relies on adequate plasma and brain concentrations to produce the required pharmacodynamic response without toxic adverse effects. In this chapter, we describe clinically significant drug-drug interactions caused by changes in pharmacokinetics (induction or inhibition of metabolism) and pharmacodynamics (inhibited, additive or synergistic activity at the drug target) of the anti-addiction agents that alter treatment success, cause harmful effects and/or have led to recommendations from regulatory bodies regarding dosage adjustments or additional monitoring for patient safety. Clinicians need to be aware of the potential for these drug-drug interactions and prescribe and monitor patients for safety and efficacy as needed.

A quantitative, peripherally accessible biomarker for neuropathic pain has great potential to improve clinical outcomes. Based on the premise that peripheral and central immunity contribute to neuropathic pain mechanisms, we hypothesized that biomarkers could be identified from the whole blood of adult male rats, by integrating graded chronic constriction injury (CCI), ipsilateral lumbar dorsal quadrant (iLDQ) and whole blood transcriptomes, and pathway analysis with pain behavior. Correlational bioinformatics identified a range of putative biomarker genes for allodynia intensity, many encoding for proteins with a recognized role in immune/nociceptive mechanisms. A selection of these genes was validated in a separate replication study. Pathway analysis of the iLDQ transcriptome identified Fcγ and Fce signaling pathways, among others. This study is the first to employ the whole blood transcriptome to identify pain biomarker panels. The novel correlational bioinformatics, developed here, selected such putative biomarkers based on a correlation with pain behavior and formation of signaling pathways with iLDQ genes. Future studies may demonstrate the predictive ability of these biomarker genes across other models and additional variables.

The TaqI A polymorphism (A(1)) of the dopamine D(2) receptor gene (DRD2), although not a specific predictor of opioid dependence, has been strongly associated with high levels of prior heroin use and poor treatment outcomes among methadone maintenance patients. The aims of this study were to confirm these findings via a retrospective analysis of A(1) allele frequency in methadone (n = 46) and buprenorphine (n = 25) patients, and non-opioid-dependent controls (n = 95). Subjects were genotyped at the DRD2 TaqI A locus using PCR amplification followed by TaqI restriction enzyme digestion and gel electrophoresis. For methadone and buprenorphine subjects, heroin use (prior to treatment), treatment outcomes, and withdrawal occurrence were determined from comprehensive case notes. No significant differences in A(1) allele frequency (%) were observed between: methadone (19.6%), buprenorphine (18.0%), and control (17.9%) groups (P > 0.7); successful and poor treatment outcome groups, methadone: 20.0% and 19.2%, respectively (P = 1.0); buprenorphine: 18.4% and 20.0%, respectively (P = 1.0). Also, there were no significant relationships between TaqI A genotype and prior heroin use (P = 0.47). However, among the successful methadone subjects, significantly fewer A(1) allele carriers experienced withdrawal than non-A(1) carriers (P = 0.04). In conclusion, the DRD2 genotype effects did not affect opioid maintenance treatment outcomes. This suggests the need for a further prospective investigation into the role of the DRD2 A(1) allele in heroin use and response to maintenance pharmacotherapies for opioid dependence.

For opioids requiring CYP2D6 O-demethylation to active metabolites, poor metabolizers have reduced metabolite formation and minimal pain reduction. Clinically, this has only reliably been shown for tramadol. Ultra-rapid metabolizers have an increased risk of toxicity especially for codeine. ABCB1 genetics show no consistent findings. In Asian populations, the high OPRM1 118A>G frequency associates with higher opioid dosage requirements. Clinical translation of opioid genetics is premature because many important pain and addiction phenotype factors contribute.

Methadone is the major opioid substitution therapy for opioid dependence. Dosage is highly variable and is often controlled by the patient and prescriber according to local and national policy and guidelines. Nevertheless many genetic factors have been investigated including those affecting its metabolism (CYP2B6-consistent results), efflux transport (P-gp-inconsistent results), target μ-opioid receptor (μ-opioid receptor-inconsistent results) and a host of other receptors (DRD2) and signaling elements (GIRK2 and ARRB2; not replicated). None by themselves have been able to substantially explain dosage variation (the major but not sole end point). When multiple genes have been combined such as ABCB1, CYP2B6, OPRM1 and DRD2 a greater contribution to dosage variation was found but not as yet replicated. As stabilization of dosage needs to be made rapidly, it is imperative that larger internationally based studies be instigated so that genetic contribution to dosage can be properly assessed, which may or may not tailor to different ethnic groups and each country’s policy towards an outcome that benefits all.

This study aimed to investigate whether CYP3A4/5 genetic variants, together with clinical and patient factors, influence serum fentanyl and norfentanyl concentrations and their ratio in cancer pain patients receiving transdermal fentanyl.CYP3A4*22 and CYP3A5*3 polymorphisms were analysed in 620 cancer pain patients receiving transdermal fentanyl (12.5-700 μg/h) from the European Pharmacogenetic Opioid Study. Using stepwise linear regression, CYP3A4/5 genetic variability was examined in combination with patient factors relating to organ drug elimination function and ABCB1 genetics for their association with serum fentanyl and norfentanyl concentrations and metabolic ratio (MR) (norfentanyl : fentanyl).Delivery rate-adjusted serum fentanyl concentrations (0.0012-1.1 nmol/l/μg.h) and MRs (0.08-499) varied widely. Only 43% of variability in serum fentanyl concentrations was accounted for by delivery rate and less than 50% by CYP3A4/5 genotypes and clinical variables (delivery rate, sex, comedications, kidney disease, BMI, serum albumin). CYP3A4*22 and CYP3A5*3 variants, CYP3A inhibitors and variables relating to liver and kidney function (serum albumin, glomerular filtration rate, kidney disease, BMI) were associated with MR, but accounted for only 14% of variability.Serum fentanyl concentrations and MR vary considerably between cancer pain patients on transdermal fentanyl patches. CYP3A4*22 and CYP3A5*3 genotypes, and multiple clinical factors, combine to influence transdermal fentanyl pharmacokinetics, but accounted for only a small proportion of variability in this study. Identification of the remaining factors determining serum fentanyl concentrations, and their relationship to efficacy and adverse effects may aid in improving the safety and effectiveness of transdermal fentanyl.

A quantitative, peripherally accessible biomarker for neuropathic pain has great potential to improve clinical outcomes. Based on the premise that peripheral and central immunity contribute to neuropathic pain mechanisms, we hypothesized that biomarkers could be identified from the whole blood of adult male rats, by integrating graded chronic constriction injury (CCI), ipsilateral lumbar dorsal quadrant (iLDQ) and whole blood transcriptomes, and pathway analysis with pain behavior. Correlational bioinformatics identified a range of putative biomarker genes for allodynia intensity, many encoding for proteins with a recognized role in immune/nociceptive mechanisms. A selection of these genes was validated in a separate replication study. Pathway analysis of the iLDQ transcriptome identified Fcγ and Fcε signaling pathways, among others. This study is the first to employ the whole blood transcriptome to identify pain biomarker panels. The novel correlational bioinformatics, developed here, selected such putative biomarkers based on a correlation with pain behavior and formation of signaling pathways with iLDQ genes. Future studies may demonstrate the predictive ability of these biomarker genes across other models and additional variables.

Opioids are used for acute and chronic pain and dependency. They have a narrow therapeutic index and large interpatient variability in response. Genetic factors regulating their pharmacokinetics (metabolizing enzymes, transporters) and pharmacodynamics (receptors and signal transduction elements) are contributors to such variability. The polymorphic CYP2D6 regulates the O-demethylation of codeine and other weak opioids to more potent metabolites with poor metabolizers having reduced antinociception in some cases. Some opioids are P-glycoprotein substrates, whereas, ABCB1 genotypes inconsistently influence opioid pharmacodynamics and dosage requirements. Single-nucleotide polymorphisms in the mu opioid receptor gene are associated with increasing morphine, but not methadone dosage requirements and altered efficacy of mu opioid agonists and antagonists. As knowledge regarding the interplay between genes affecting opioid pharmacokinetics including cerebral kinetics and pharmacodynamics increases, our understanding of the role of pharmacogenomics in mediating interpatient variability in efficacy and side effects to this important class of drugs will be better informed. Opioid drugs as a group have withstood the test of time in their ability to attenuate acute and chronic pain. Since the isolation of morphine in the early 1800s by Friedrich Serturner, a large number of opioid drugs beginning with modification of the 4,5-epoxymorphinan ring structure were developed in order to improve their therapeutic margin, including reducing dependence and tolerance, ultimately without success.

Background and Objectives: The most common treatment for opioid dependence is substitution therapy with another opioid such as methadone. The methadone dosage is individualized but highly variable, and program retention rates are low due in part to nonoptimal dosing resulting in withdrawal symptoms and further heroin craving and use. Methadone is a substrate for the P-glycoprotein transporter, encoded by the ABCB1 gene, which regulates central nervous system exposure. This retrospective study aimed to investigate the influence of ABCB1 genetic variability on methadone dose requirements. Methods: Genomic deoxyribonucleic acid was isolated from opioid-dependent subjects (n 60) and non– opioid-dependent control subjects (n 60), and polymerase chain reaction–restriction fragment length polymorphism and allele-specific polymerase chain reaction were used to determine the presence of single nucleotide polymorphisms at positions 61, 1199, 1236, 2677, and 3435. ABCB1 haplotypes were inferred with PHASE software (version 2.1). Results: There were no significant differences in the allele or genotype frequencies of the individual single nucleotide polymorphisms or haplotypes between the 2 populations. ABCB1 genetic variability influenced daily methadone dose requirements, such that subjects carrying 2 copies of the wild-type haplotype required higher doses compared with those with 1 copy and those with no copies (98.3 10.4, 58.6 20.9, and 55.4 26.1 mg/d, respectively; P .029). In addition, carriers of the AGCTT haplotype required significantly lower doses than noncarriers (38.0 16.8 and 61.3 24.6 mg/d, respectively; P .04). Conclusion: Although ABCB1 genetic variability is not related to the development of opioid dependence, identification of variant haplotypes may, after larger prospective studies have been performed, provide clinicians with a tool for methadone dosage individualization. (Clin Pharmacol Ther 2006;80:682-90.)

Daniel T Barratt1, Janet K Coller1, Richard Hallinan2, Andrew Byrne2, Jason M White1, David JR Foster3, Andrew A Somogyi1,41Discipline of Pharmacology, School of Medical Sciences, University of Adelaide, Adelaide, South Australia; 2The Byrne Surgery, Specialist Drug and Alcohol Practice, Redfern, New South Wales; 3Division of Health Sciences, Sansom Institute, School of Pharmacy and Medical Sciences, University of South Australia, Adelaide, South Australia; 4Department of Clinical Pharmacology, Royal Adelaide Hospital, Adelaide, South Australia, AustraliaBackground: Genetic variability in ABCB1, encoding the P-glycoprotein efflux transporter, has been linked to altered methadone maintenance treatment dose requirements. However, subsequent studies have indicated that additional environmental or genetic factors may confound ABCB1 pharmacogenetics in different methadone maintenance treatment settings. There is evidence that genetic variability in OPRM1, encoding the mu opioid receptor, and ABCB1 may interact to affect morphine response in opposite ways. This study aimed to examine whether a similar gene-gene interaction occurs for methadone in methadone maintenance treatment.Methods: Opioid-dependent subjects (n = 119) maintained on methadone (15–300 mg/day) were genotyped for five single nucleotide polymorphisms of ABCB1 (61A > G; 1199G > A; 1236C > T; 2677G > T; 3435C > T), as well as for the OPRM1 18A > G single nucleotide polymorphism. Subjects’ methadone doses and trough plasma (R)-methadone concentrations (Ctrough) were compared between ABCB1 haplotypes (with and without controlling for OPRM1 genotype), and between OPRM1 genotypes (with and without controlling for ABCB1 haplotype).Results: Among wild-type OPRM1 subjects, an ABCB1 variant haplotype group (subjects with a wild-type and 61A:1199G:1236C:2677T:3435T haplotype combination, or homozygous for the 61A:1199G:1236C:2677T:3435T haplotype) had significantly lower doses (median ± standard deviation 35 ± 5 versus 180 ± 65 mg/day, P < 0.01) and Ctrough (78 ± 22 versus 177 ± 97 ng/mL, P < 0.05) than ABCB1 wild-type subjects. Among subjects with the most common ABCB1 haplotype combination (wild-type with 61A:1199G:1236T:2677T:3435T), the OPRM1 118 A/G genotype was associated with a significantly higher Ctrough than 118 A/A (250 ± 126 versus 108 ± 36 ng/mL, P = 0.016). No ABCB1 haplotype group or OPRM1 genotype was associated with dose or Ctrough without taking into account confounding genetic variability at the other locus. Therefore, two interacting pharmacogenetic determinants of methadone maintenance treatment response were identified, ie, ABCB1, where variants are associated with lower methadone requirements, and OPRM1, where the variant is associated with higher methadone requirements.Conclusion: These opposing pharmacogenetic effects therefore need to be considered in combination when assessing methadone maintenance treatment pharmacogenetics.Keywords: methadone, opiate substitution treatment, ABCB1, P-glycoprotein, OPRM1, receptors, opioid, mu