Vasopressor requirement during spinal anaesthesia for caesarean delivery

Numerous clinical trials have evaluated the response to vasopressors to prevent and or treat hypotension during spinal anaesthesia for elective caeserean delivery. For decades, ephedrine has been considered a safe option based on classic studies in sheep that suggested deleterious effects of pure α-adrenergic agonists on uteroplacental blood flow. Ephedrine has been widely used in a variety of regimens (different bolus doses, infusions and in combination with phenylephrine) although no consensus has ever been achieved as to which of these modes of administration provides the most reliable and effective response. Ephedrine is a sympathomimetic amine, the principal mechanism of its action relies on its direct and indirect actions on the adrenergic receptor system (both an α- and β-adrenergic agonist).

Several single nucleotide polymorphisms (SNPs) that have been described in the gene encoding the human β ₂ -adrenergic receptor (β ₂ AR) affect the function of the receptor in vitro . Substitution of glycine for arginine at residue 16 (Arg16Gly) has been associated with enhanced agonist-induced desensitization, while substitution of glutamic acid for glutamine at position 27 (Gln27Glu) has been associated with resistance to desensitization. Significant differences in the response of individuals to β ₂ AR therapeutic manipulation related to the particular genotype/haplotype of the β ₂ AR have been demonstrated.

A pharmacogenetic study in an obstetric population showed that the incidence and severity of maternal hypotension after spinal anaesthesia for caesarean delivery and the response to treatment is clearly affected by β ₂ AR genotype/haplotype. Women Gly16 homozygous, carrying one or two Glu at position 27 (heterozygous or homozygous for the Glu27 variant) were found to require significantly less vasopressors (ephedrine) for treatment of hypotension during spinal anaesthesia. The two haplotypes that seem to ‘protect’ women from requiring higher doses of ephedrine are relatively common in Caucasians, and in this study 20% of the women carried either one of these haplotypes. This pharmacogenetic effect may explain in part why the numerous studies trying to prevent or treat hypotension during spinal anaesthesia for caesarean section failed to define one single optimal strategy (fluid loading, ephedrine or phenylephrine) that would ‘fit all’.

Since the incidence of spinal hypotension and vasopressor use is reduced in preeclampsia , we further hypothesized that haplotypes of β ₂ AR gene influence haemodynamics during spinal anaesthesia for caesarean delivery in women diagnosed with severe pre-eclampsia. In a prospective case-control study, we compared the incidence of hypotension and vasopressor requirements in a predominantly African-American cohort. Despite a trend towards fewer pre-eclamptic women requiring vasopressors, the total vasopressor dose was higher in those in whom treatment was indicated, which was contrary to our expectations. However, no woman in the pre-eclamptic group carried the Gly16Gly/Glu27Glu haplotype, and since this was one of the two haplotypes that predicted less vasopressor requirement in normotensive women , this might provide an explanation for our unexpected results. Whether our findings are specific to African-American women remains to be determined in larger studies in all ethnic groups. These findings illustrate the importance of ethnicity when assessing genetic associations, and similar interactions between ethnicity and genetics have been suggested for other SNPs presented in this review (β ₁ AR, μ-OR). In the long term, if these findings are confirmed, clinical implications could involve using haplotype of β ₂ AR to predict spinal hypotension and to guide haemodynamic management in women with compromised cardiovascular function and altered uteroplacental perfusion.

Meanwhile, the effects of ephedrine on the foetus have been revisited recently. Evidence that ephedrine crosses the placenta to a greater extent and undergoes less early metabolism and redistribution than phenylephrine (a direct α-adrenergic agonist) causing direct foetal metabolic acidosis has made ephedrine less desirable as a first-line treatment. The proposed mechanism is that direct foetal β-adrenergic stimulation increases anaerobic glycolysis and causes a hypermetabolic state. It is possible that the response of the foetus to β-adrenergic manipulation may also be genetically mediated and that foetal haplotype may impact on the response to ephedrine given to the mother. This hypothesis is currently under investigation and may provide some insight on foetal acidosis and metabolic responses in neonates born to mothers who have received β-agonists (ephedrine and/or other β-stimulants prescribed for tocolysis or bronchodilation) prior to delivery.

β-blockers and management of cardiovascular disease and hypertension

β-adrenergic receptors (β-AR) mediate chronotropic and inotropic responses to catecholamines, which is of particular interest to anaesthesiologists. β-blockade is recommended to treat hypertensive disorders and provide myocardial protection either peri-operatively or after acute myocardial infarction (MI). Indeed administration of β-blockers after an acute ischemic event has become a quality measure by which hospitals are judged.

In a large prospective cohort study in patients receiving β-blockers after an acute coronary event, four common β ₁ and β ₂ AR SNPs were examined. Increases in mortality rates were found with possession of certain variants in the β ₂ AR, rising to 20% mortality at 3 years according to the haplotype combination of Arg16Gly and Gln27Gly. It appears from this study that patients with variants impairing β ₂ AR downregulation (Gly16/Glu27), where receptor function does not undergo desensitization, benefit from β-blocker therapy. Conversely, those with genotypes enhancing downregulation (Arg16/Gln27) do not benefit from β-blockers, most likely because less receptor is present at the cell surface, which mimics βAR antagonist activity. In fact, the administration of β-blockers to such patients appears to unmask negative effects. Also of interest, the authors report no association of the β ₁ AR variants with mortality regardless of β-blocker therapy. Pending replication, this study provides compelling evidence that genetic variability of the β ₂ AR has direct clinical relevance.

A common polymorphism of β ₁ AR, Arg389Gly, has been extensively evaluated both in vitro and in clinical studies. The Arg389 variant been shown in vitro to display a greater response to agonist stimulation, and individuals with the Gly389 allele have a decreased response to β-blockade.

Based on recently accumulated evidence in the literature, a large randomized clinical trial (INVEST-GENES, n = 5,892 hypertensive patients with coronary artery disease) tested the hypothesis that patients with a combination of the Ser49/Arg389 haplotype of β ₁ AR or a combination of the Arg16/Gln27 haplotype of β ₂ AR would be at relatively higher risk for cardiovascular events and that atenolol would be more beneficial as compared with a calcium-channel antagonist (verapamil sustained release) in patients carrying the Ser49/Arg389 β ₁ AR haplotype. The increase in mortality risk among patients with one or two copies of the Ser49/Arg389 haplotype was highly significant in patients randomly assigned to verapamil SR but not in patients assigned to atenolol. These findings suggest that atenolol offsets the mortality risk associated with the β ₁ AR haplotype, consistent with prior observations that patients bearing the wild-type alleles are more responsive to β-blocker therapy in settings of blood pressure lowering, improvement in ejection fraction, and survival in heart failure. Findings related to β ₂ AR haplotype did not allow to draw any firm associations and given the inconsistencies in the literature, these findings require independent replication. The authors concluded that β ₁ AR haplotype variation is associated with mortality risk, and β-blockers may be preferred in subgroups of patients defined by adrenergic receptor polymorphisms.

Of interest, the Gly389 allele is more frequent in African-Americans (42%) compared with Caucasians (28%) and this has been suggested to in part explain the decreased sensitivity to β-blockade reported in African-Americans. This specific question has since then been reevaluated, and may be of particular interest for obstetric anaesthesiologists and obstetricians involved with management of hypertension in pregnancy and pre-eclampsia. One study specifically sought to determine whether ethnic differences in response to β-blockade could be attributed to distribution of genetic variants in the β ₁ AR. Exercise-induced tachycardia was evaluated before and after atenolol response, and atenolol resulted in a greater reduction in exercise heart rate in whites than in blacks. β ₁ AR Arg389 was independently associated with a greater reduction in heart rate, however ethnic differences in response to atenolol remained significant after adjustment for genotype. Therefore, ethnic differences in sensitivity to atenolol persist even after accounting for different distributions of genetic variants of β ₁ AR, suggesting that additional factors, yet to be identified, most likely contribute to these ethnic differences. Another study suggested that β ₁ AR genotype contributes significantly in the observed differences in β-blocker response between Caucasians and African-Americans patients with heart failure. Taken together, these results suggest that treatment response should take into account both ethnicity and genotype. A recent review summarizes the effect of relevant polymorphisms affecting pharmacodynamics and pharmacokinetics of medication prescribed for the management of cardiovascular disease.

Further evaluation of the effect of βAR haplotypes in the context of hypertensive management in pregnancy will be of interest. There has been to date no clinical trial examining the response to β-blockers in pre-eclamptic women according to genotype of β ₁ AR, which may provide in part an explanation for the observed ineffectiveness of labetalol in some hypertensive/pre-eclamptic women.

Bronchodilators for management of asthma

There have been numerous studies assessing the association between asthma phenotypes and genetic variability of β ₂ AR. A meta-analysis concluded that neither the Gly16 nor the Glu27 allele contributes to asthma susceptibility overall or to bronchial hyper-responsiveness. Gly16 homozygotes however did have a much higher risk for nocturnal asthma and asthma severity than Arg16 homozygotes. Therefore, SNPs of β ₂ AR gene are not major risk factors for the development of asthma, rather they are ‘disease modifiers’ and are important in determining drug response (pharmacogenetic effect).

Initial studies described an enhanced response to β ₂ -agonist bronchodilators in asthmatic subjects homozygous for the wild-type allele (Arg16) compared to patients homozygous for Gly16, which would appear to be consistent with the prediction of the in vitro findings that demonstrated increased receptor down-regulation in presence of the Gly16 variant. However, recent genotype-stratified studies on treatment outcome in patients with mild asthma determined that patients homozygous for Gly16 improved in the long term with albuterol or long-acting β ₂ -agonists while those homozygous for Arg16 did not . With several studies showing that asthmatic patients who are Arg16 homozygous do not benefit from short-acting β-agonists in either the absence or presence of concurrent inhaled corticosteroid use, investigation of alternate treatment strategies that may help this group is on its way. Pharmacogenetics should influence the clinical management of asthma in a very near future.

Tocolytics for management of preterm labour and delivery

Stimulation of the β ₂ AR results in uterine smooth muscle relaxation, and thus the β ₂ AR has long been a therapeutic target for the treatment of preterm labour. β ₂ -agonist therapy, in common with virtually all tocolytics, has not been consistently successful at stopping preterm labour or prolonging pregnancy, in part due to the multifactorial nature of preterm labour, and possibly because of a wide variability in therapeutic response within the population. The mechanisms involved in regulation of myometrial smooth muscle contraction and relaxation in preterm labour or even at term are not yet fully elucidated. Polymorphisms of β ₂ AR (Arg16Gly and Gln27Glu) have been associated in several studies with preterm labour and delivery. Arg16 homozygosity of β ₂ AR genotype appears to confer a protective effect against preterm delivery while the Glu27 variant might increase the risk for preterm delivery. Furthermore, we have shown a pharmacogenetic effect, with a better response to β ₂ agonist therapy (hexoprenaline) for tocolysis in women Arg16 homozygous with idiopathic preterm labour between 24 and 34 weeks gestation. This had a significant impact on neonatal outcomes, with higher birth weights and less neonatal ICU admissions for respiratory or other complications due to prematurity in babies born to mothers with that genotype. It remains to be determined whether β ₂ AR genotype influences the severity of the disease (i.e. that Arg16 homozygote women present with a milder disease than women with other genotypes) or directly affects the response to therapy. Meanwhile, a variety of genomic studies have examined the influence of genetic variants on the incidence of preterm labour, and proteomic studies to validate biomarkers that could identify women at risk for preterm delivery and serve as predictive tools is ongoing.

Analgesia and pain-related candidate genes

Interindividual variability in pain perception and sensitivity to analgesic therapy with a large unpredictability in efficacy, side effects and tolerance profiles to opioids is well described. Numerous candidate genes have been considered as suitable targets for the study of the genetic basis of pain and or analgesia. In addition, a genetic database of knock-out mice allowing the study of genetic variations in the context of specific pain phenotypes was made public.

Recently an extremely rare phenotype characterized by a total absence of pain perception (‘congenital indifference to pain’) with no associated neuropathy has been associated with the mutations in the gene SCN9A, encoding the α-subunit of the voltage-gated sodium channel, Na _v 1.7. Individuals with loss-of-function mutations of the Na _V 1.7 lack protective mechanisms that allow tissue damage detection and suffer severe injuries because they do not learn pain-avoiding behaviors. This discovery opens new directions for development of novel generations of drugs with blocking Na _v 1.7 proprieties, which should provide more selective and safe analgesia. Meanwhile, we are still in the era of opioid therapy, and the analgesic effect may be influenced by alterations in the metabolism of analgesic drugs (cytochrome P450), variants coding for the μ-opioid receptor (μOR) as well as other targets.

The codeine ‘story’

Cytochrome P450 (CYP450) is a super-family of liver enzymes that catalyze phase 1 drug metabolism. The gene coding for this enzyme was found to be highly polymorphic, with more than 75 different CYP2D6 alleles ( http://www.imm.ki.se/cypalleles ), resulting in a variable enzymatic activity ranging from 1 to 200%. As a result, each individual can be classified as having an “ultra-rapid metabolism”, an “extensive metabolism”, a “normal metabolism” or a “poor metabolism” and microarray technology is available to classify individuals according to their metabolic phenotype. Of note, CYP2D6 activity is absent in approximately 7–10% Caucasians. Codeine is a pro-drug and needs be converted into morphine to elicit its analgesic effect, therefore ‘poor metabolizers do not achieve analgesia with codeine while they do encounter side effects such as nausea and vomiting. Conversely, codeine intoxication can be anticipated with ultra-rapid CYP2D6 metabolism.

While codeine is undoubtedly not a wonder analgesic, it is still prescribed because of the belief that being a weak opioid, it is safe. There has been a recent FDA warning on codeine use in nursing mothers ( http://www.fda.gov/bbs/topics/NEWS/2007/NEW01685.html ) following the death of a breastfed 13 days old neonate thought to have suffered a morphine overdose because his mother was taking codeine and was a CYP2D6 ultra-rapid metabolizer. Since that fatal report in 2006, numerous publications have addressed the issue of potential neonatal and pediatric codeine intoxications according to pharmacokinetic phenotype. This case exemplifies very precisely the irrefutable input of diagnostic pharmacogenetic testing and carries direct clinical implications for obstetric anaesthesiologists, obstetricians and neonatologists.

Similarly, the effect of tramadol, a centrally acting μ-opioid agonist that has a dual mechanism of action, binding to the μORs and weakly inhibiting the neuronal reuptake of norepinephrine and serotonin, is also influenced by the CYP2D6 genotype and interactions with any other co-administered medication undergoing CYP2D6 metabolism should be anticipated.

Neuraxial labour analgesia and post-caesarean analgesia

The μOR, encoded by genetic locus OPRM1 , has been the focus of several genetic studies because this receptor is the primary site of action for many endogenous opioid peptides and the major target for opioid analgesics, such as morphine, heroin (diamorphine), fentanyl, and methadone. The μOR is a GPCR, and a number of SNPs have been described for OPRM1 . At nucleotide position 118, an adenine substitution by a guanine (A118 G), has been reported to occur with an allelic frequency of 10–30% among Caucasians with a higher prevalence among Asians. The major interest for this particular SNP is due to its pharmacological and physiological consequences. In vitro , the presence of at least one G118 allele has been shown to increase the binding affinity and potency of β-endorphin. Thus, individuals carrying the variant receptor gene could show differences in some of the functions mediated by β-endorphin action at the altered μOR, such as higher thresholds to pain. Consistent with this laboratory finding, one in vivo study in a human experimental pain model demonstrated that volunteers carrying a G118 allele exhibited lower sensitivity to pressure pain compared with A118 homozygotes. The exact relationship of this polymorphism with the responses to oral opioids for treatment of chronic pain, post-operative requirements of intravenous alfentanil or morphine or the adverse effects and toxicity profiles appear to go ‘in the opposite direction’. Individuals carrying the variant G118 allele may actually require higher doses of morphine, which is in contrast with what is expected if the variant μOR is truly associated with an increased endogenous opioid activity.

Our group has demonstrated that the A118 G polymorphism impacts on the median effective dose (ED50) of intrathecal fentanyl in labour; nulliparous women carrying the variant G118 allele required substantially lower doses of intrathecal fentanyl to achieve adequate analgesia early in labour (with a 1.5 to 2 fold difference between genetic groups). Of note, cervical dilatation at the time of analgesia request was significantly less in women homozygous for the wild-type variant (A118) than that in women carrying one or two variant alleles (G118). This is of interest, since women received combined-spinal epidural (CSE) analgesia by their own request, at the time they experienced painful contractions. Since, it has been demonstrated previously that epidural analgesic requirements increase with progress of labour and cervical dilatation, the expectation would be that women carrying the mutant G allele should have greater analgesic requirements due to the greater cervical dilatation at which they requested analgesia; our finding that these women require less fentanyl therefore may actually underestimate the true effect of genotype. This is consistent with the premise that women carrying the G allele may have higher pain tolerance and therefore request epidural analgesia at a later stage in labour. Our finding of a significant difference in the median effective dose (ED50) according to genotype is clinically relevant, because provision of optimal labour analgesia remains an ongoing challenge for obstetric anaesthesiologists. A major goal of labour analgesia is minimal motor impairment, emphasizing the need to minimize local anaesthetic use. Similarly, there is a need to reduce opioid doses in order to minimize opioid-related side effects such as pruritus and foetal bradycardia (which appear to be dose-dependant). Therefore, according to our findings, genotyping may well help improve the delivery of labour analgesia because a 1.5–2 fold increase in fentanyl dose is not trivial.

We have also evaluated the effect of the A118 G polymorphism on the duration of intrathecal fentanyl analgesia in early labour and did not find any difference between genotypic groups. While the A118 G polymorphism may influence drug potency, there may be no pharmacokinetic effect altering duration of drug action.

As mentioned above, our findings in labouring women are in disagreement with studies looking at post-operative intravenous consumption of morphine after general or orthopedic surgery. In addition, three studies specifically evaluated post-caesarean analgesia with intrathecal or intravenous morphine. One study did not find a difference in the duration of intrathecal morphine analgesia or need for analgesic supplementation in women carrying the G118 allele; however, among these women, pruritus was less frequent during the first 24 h post-operatively. The two other studies, both from Singapore, reported that Asian women with at least one G118 allele exhibited increased post-caesarean morphine consumption with intravenous patient-controlled analgesia.

These differences may be due to the fact that spinal and systemic opioid pharmacokinetics and pharmacodynamics may be different; enhanced analgesia in response to intrathecal fentanyl in the presence of the G118 allele may not exist in response to intravenous fentanyl or other opioids via the intravenous route. Or one could speculate that human spinal cord receptor function and signal transduction is selectively more altered by the G118 variant than supraspinal receptors. Another potential explanation and factor to bear in mind is the different nature of the nociceptive stimulus in labour versus other painful syndromes. Indeed, similar disparate results in human genetic studies of pain sensitivity have been shown to occur with other polymorphisms commonly assessed in pain studies. A polymorphism of the catechol- O -methyltransferase gene (Val158Met) has been inconsistently found to be associated with altered nociception and response to analgesic; while one study found a significant genetic association, another did not, a third described a significant effect but not for that polymorphism, and a fourth reported that findings depended on stimulus modality.

This illustrates the challenges in evaluating a genotype-phenotype association when the underlying genetic susceptibility is clearly polygenic and the phenotype is complex in itself (multifactorial, subjective, involving pain perception and/or response to analgesics).

Last, a relatively unexpected candidate gene has been suggested to modulate the perception of pain in labouring women in an ongoing study. Women who were Gly16 homozygous for the β ₂ AR reported less pain in early labour compared to others. Whether haplotypes of β ₂ AR predict more rapid and less painful labour remains to be confirmed.

Undeniably, numerous candidate genes as well as elaborate models have been suggested for the study of the genetic component of pain and several excellent reviews on the pharmacogenetics of opioids and analgesia were recently published. Nonetheless, due to the inherent complexity of studying pain (different nociceptive modalities, gender differences, limitations in extrapolating data from animal models to the response in humans, interethnic and environmental differences) in addition to the obvious polygenic nature of this, it is the design and execution of large clinical studies analyzing multiples haplotypes simultaneously that remains to be the true challenge to date. Meanwhile, genome wide association studies (GWAS) in the context of acute post-operative pain are published and researchers are actively working on gene therapies for pain. It will also be of interest to see the new insights and developments brought by more research on the SCN9A gene in the near future.

Vasopressor requirement during spinal anaesthesia for caesarean delivery

Numerous clinical trials have evaluated the response to vasopressors to prevent and or treat hypotension during spinal anaesthesia for elective caeserean delivery. For decades, ephedrine has been considered a safe option based on classic studies in sheep that suggested deleterious effects of pure α-adrenergic agonists on uteroplacental blood flow. Ephedrine has been widely used in a variety of regimens (different bolus doses, infusions and in combination with phenylephrine) although no consensus has ever been achieved as to which of these modes of administration provides the most reliable and effective response. Ephedrine is a sympathomimetic amine, the principal mechanism of its action relies on its direct and indirect actions on the adrenergic receptor system (both an α- and β-adrenergic agonist).

Several single nucleotide polymorphisms (SNPs) that have been described in the gene encoding the human β ₂ -adrenergic receptor (β ₂ AR) affect the function of the receptor in vitro . Substitution of glycine for arginine at residue 16 (Arg16Gly) has been associated with enhanced agonist-induced desensitization, while substitution of glutamic acid for glutamine at position 27 (Gln27Glu) has been associated with resistance to desensitization. Significant differences in the response of individuals to β ₂ AR therapeutic manipulation related to the particular genotype/haplotype of the β ₂ AR have been demonstrated.

A pharmacogenetic study in an obstetric population showed that the incidence and severity of maternal hypotension after spinal anaesthesia for caesarean delivery and the response to treatment is clearly affected by β ₂ AR genotype/haplotype. Women Gly16 homozygous, carrying one or two Glu at position 27 (heterozygous or homozygous for the Glu27 variant) were found to require significantly less vasopressors (ephedrine) for treatment of hypotension during spinal anaesthesia. The two haplotypes that seem to ‘protect’ women from requiring higher doses of ephedrine are relatively common in Caucasians, and in this study 20% of the women carried either one of these haplotypes. This pharmacogenetic effect may explain in part why the numerous studies trying to prevent or treat hypotension during spinal anaesthesia for caesarean section failed to define one single optimal strategy (fluid loading, ephedrine or phenylephrine) that would ‘fit all’.

Since the incidence of spinal hypotension and vasopressor use is reduced in preeclampsia , we further hypothesized that haplotypes of β ₂ AR gene influence haemodynamics during spinal anaesthesia for caesarean delivery in women diagnosed with severe pre-eclampsia. In a prospective case-control study, we compared the incidence of hypotension and vasopressor requirements in a predominantly African-American cohort. Despite a trend towards fewer pre-eclamptic women requiring vasopressors, the total vasopressor dose was higher in those in whom treatment was indicated, which was contrary to our expectations. However, no woman in the pre-eclamptic group carried the Gly16Gly/Glu27Glu haplotype, and since this was one of the two haplotypes that predicted less vasopressor requirement in normotensive women , this might provide an explanation for our unexpected results. Whether our findings are specific to African-American women remains to be determined in larger studies in all ethnic groups. These findings illustrate the importance of ethnicity when assessing genetic associations, and similar interactions between ethnicity and genetics have been suggested for other SNPs presented in this review (β ₁ AR, μ-OR). In the long term, if these findings are confirmed, clinical implications could involve using haplotype of β ₂ AR to predict spinal hypotension and to guide haemodynamic management in women with compromised cardiovascular function and altered uteroplacental perfusion.

Meanwhile, the effects of ephedrine on the foetus have been revisited recently. Evidence that ephedrine crosses the placenta to a greater extent and undergoes less early metabolism and redistribution than phenylephrine (a direct α-adrenergic agonist) causing direct foetal metabolic acidosis has made ephedrine less desirable as a first-line treatment. The proposed mechanism is that direct foetal β-adrenergic stimulation increases anaerobic glycolysis and causes a hypermetabolic state. It is possible that the response of the foetus to β-adrenergic manipulation may also be genetically mediated and that foetal haplotype may impact on the response to ephedrine given to the mother. This hypothesis is currently under investigation and may provide some insight on foetal acidosis and metabolic responses in neonates born to mothers who have received β-agonists (ephedrine and/or other β-stimulants prescribed for tocolysis or bronchodilation) prior to delivery.

β-blockers and management of cardiovascular disease and hypertension

β-adrenergic receptors (β-AR) mediate chronotropic and inotropic responses to catecholamines, which is of particular interest to anaesthesiologists. β-blockade is recommended to treat hypertensive disorders and provide myocardial protection either peri-operatively or after acute myocardial infarction (MI). Indeed administration of β-blockers after an acute ischemic event has become a quality measure by which hospitals are judged.

In a large prospective cohort study in patients receiving β-blockers after an acute coronary event, four common β ₁ and β ₂ AR SNPs were examined. Increases in mortality rates were found with possession of certain variants in the β ₂ AR, rising to 20% mortality at 3 years according to the haplotype combination of Arg16Gly and Gln27Gly. It appears from this study that patients with variants impairing β ₂ AR downregulation (Gly16/Glu27), where receptor function does not undergo desensitization, benefit from β-blocker therapy. Conversely, those with genotypes enhancing downregulation (Arg16/Gln27) do not benefit from β-blockers, most likely because less receptor is present at the cell surface, which mimics βAR antagonist activity. In fact, the administration of β-blockers to such patients appears to unmask negative effects. Also of interest, the authors report no association of the β ₁ AR variants with mortality regardless of β-blocker therapy. Pending replication, this study provides compelling evidence that genetic variability of the β ₂ AR has direct clinical relevance.

A common polymorphism of β ₁ AR, Arg389Gly, has been extensively evaluated both in vitro and in clinical studies. The Arg389 variant been shown in vitro to display a greater response to agonist stimulation, and individuals with the Gly389 allele have a decreased response to β-blockade.

Based on recently accumulated evidence in the literature, a large randomized clinical trial (INVEST-GENES, n = 5,892 hypertensive patients with coronary artery disease) tested the hypothesis that patients with a combination of the Ser49/Arg389 haplotype of β ₁ AR or a combination of the Arg16/Gln27 haplotype of β ₂ AR would be at relatively higher risk for cardiovascular events and that atenolol would be more beneficial as compared with a calcium-channel antagonist (verapamil sustained release) in patients carrying the Ser49/Arg389 β ₁ AR haplotype. The increase in mortality risk among patients with one or two copies of the Ser49/Arg389 haplotype was highly significant in patients randomly assigned to verapamil SR but not in patients assigned to atenolol. These findings suggest that atenolol offsets the mortality risk associated with the β ₁ AR haplotype, consistent with prior observations that patients bearing the wild-type alleles are more responsive to β-blocker therapy in settings of blood pressure lowering, improvement in ejection fraction, and survival in heart failure. Findings related to β ₂ AR haplotype did not allow to draw any firm associations and given the inconsistencies in the literature, these findings require independent replication. The authors concluded that β ₁ AR haplotype variation is associated with mortality risk, and β-blockers may be preferred in subgroups of patients defined by adrenergic receptor polymorphisms.

Of interest, the Gly389 allele is more frequent in African-Americans (42%) compared with Caucasians (28%) and this has been suggested to in part explain the decreased sensitivity to β-blockade reported in African-Americans. This specific question has since then been reevaluated, and may be of particular interest for obstetric anaesthesiologists and obstetricians involved with management of hypertension in pregnancy and pre-eclampsia. One study specifically sought to determine whether ethnic differences in response to β-blockade could be attributed to distribution of genetic variants in the β ₁ AR. Exercise-induced tachycardia was evaluated before and after atenolol response, and atenolol resulted in a greater reduction in exercise heart rate in whites than in blacks. β ₁ AR Arg389 was independently associated with a greater reduction in heart rate, however ethnic differences in response to atenolol remained significant after adjustment for genotype. Therefore, ethnic differences in sensitivity to atenolol persist even after accounting for different distributions of genetic variants of β ₁ AR, suggesting that additional factors, yet to be identified, most likely contribute to these ethnic differences. Another study suggested that β ₁ AR genotype contributes significantly in the observed differences in β-blocker response between Caucasians and African-Americans patients with heart failure. Taken together, these results suggest that treatment response should take into account both ethnicity and genotype. A recent review summarizes the effect of relevant polymorphisms affecting pharmacodynamics and pharmacokinetics of medication prescribed for the management of cardiovascular disease.

Further evaluation of the effect of βAR haplotypes in the context of hypertensive management in pregnancy will be of interest. There has been to date no clinical trial examining the response to β-blockers in pre-eclamptic women according to genotype of β ₁ AR, which may provide in part an explanation for the observed ineffectiveness of labetalol in some hypertensive/pre-eclamptic women.

Bronchodilators for management of asthma

There have been numerous studies assessing the association between asthma phenotypes and genetic variability of β ₂ AR. A meta-analysis concluded that neither the Gly16 nor the Glu27 allele contributes to asthma susceptibility overall or to bronchial hyper-responsiveness. Gly16 homozygotes however did have a much higher risk for nocturnal asthma and asthma severity than Arg16 homozygotes. Therefore, SNPs of β ₂ AR gene are not major risk factors for the development of asthma, rather they are ‘disease modifiers’ and are important in determining drug response (pharmacogenetic effect).

Initial studies described an enhanced response to β ₂ -agonist bronchodilators in asthmatic subjects homozygous for the wild-type allele (Arg16) compared to patients homozygous for Gly16, which would appear to be consistent with the prediction of the in vitro findings that demonstrated increased receptor down-regulation in presence of the Gly16 variant. However, recent genotype-stratified studies on treatment outcome in patients with mild asthma determined that patients homozygous for Gly16 improved in the long term with albuterol or long-acting β ₂ -agonists while those homozygous for Arg16 did not . With several studies showing that asthmatic patients who are Arg16 homozygous do not benefit from short-acting β-agonists in either the absence or presence of concurrent inhaled corticosteroid use, investigation of alternate treatment strategies that may help this group is on its way. Pharmacogenetics should influence the clinical management of asthma in a very near future.

Tocolytics for management of preterm labour and delivery

Stimulation of the β ₂ AR results in uterine smooth muscle relaxation, and thus the β ₂ AR has long been a therapeutic target for the treatment of preterm labour. β ₂ -agonist therapy, in common with virtually all tocolytics, has not been consistently successful at stopping preterm labour or prolonging pregnancy, in part due to the multifactorial nature of preterm labour, and possibly because of a wide variability in therapeutic response within the population. The mechanisms involved in regulation of myometrial smooth muscle contraction and relaxation in preterm labour or even at term are not yet fully elucidated. Polymorphisms of β ₂ AR (Arg16Gly and Gln27Glu) have been associated in several studies with preterm labour and delivery. Arg16 homozygosity of β ₂ AR genotype appears to confer a protective effect against preterm delivery while the Glu27 variant might increase the risk for preterm delivery. Furthermore, we have shown a pharmacogenetic effect, with a better response to β ₂ agonist therapy (hexoprenaline) for tocolysis in women Arg16 homozygous with idiopathic preterm labour between 24 and 34 weeks gestation. This had a significant impact on neonatal outcomes, with higher birth weights and less neonatal ICU admissions for respiratory or other complications due to prematurity in babies born to mothers with that genotype. It remains to be determined whether β ₂ AR genotype influences the severity of the disease (i.e. that Arg16 homozygote women present with a milder disease than women with other genotypes) or directly affects the response to therapy. Meanwhile, a variety of genomic studies have examined the influence of genetic variants on the incidence of preterm labour, and proteomic studies to validate biomarkers that could identify women at risk for preterm delivery and serve as predictive tools is ongoing.

Analgesia and pain-related candidate genes

Interindividual variability in pain perception and sensitivity to analgesic therapy with a large unpredictability in efficacy, side effects and tolerance profiles to opioids is well described. Numerous candidate genes have been considered as suitable targets for the study of the genetic basis of pain and or analgesia. In addition, a genetic database of knock-out mice allowing the study of genetic variations in the context of specific pain phenotypes was made public.

Recently an extremely rare phenotype characterized by a total absence of pain perception (‘congenital indifference to pain’) with no associated neuropathy has been associated with the mutations in the gene SCN9A, encoding the α-subunit of the voltage-gated sodium channel, Na _v 1.7. Individuals with loss-of-function mutations of the Na _V 1.7 lack protective mechanisms that allow tissue damage detection and suffer severe injuries because they do not learn pain-avoiding behaviors. This discovery opens new directions for development of novel generations of drugs with blocking Na _v 1.7 proprieties, which should provide more selective and safe analgesia. Meanwhile, we are still in the era of opioid therapy, and the analgesic effect may be influenced by alterations in the metabolism of analgesic drugs (cytochrome P450), variants coding for the μ-opioid receptor (μOR) as well as other targets.

The codeine ‘story’

Cytochrome P450 (CYP450) is a super-family of liver enzymes that catalyze phase 1 drug metabolism. The gene coding for this enzyme was found to be highly polymorphic, with more than 75 different CYP2D6 alleles ( http://www.imm.ki.se/cypalleles ), resulting in a variable enzymatic activity ranging from 1 to 200%. As a result, each individual can be classified as having an “ultra-rapid metabolism”, an “extensive metabolism”, a “normal metabolism” or a “poor metabolism” and microarray technology is available to classify individuals according to their metabolic phenotype. Of note, CYP2D6 activity is absent in approximately 7–10% Caucasians. Codeine is a pro-drug and needs be converted into morphine to elicit its analgesic effect, therefore ‘poor metabolizers do not achieve analgesia with codeine while they do encounter side effects such as nausea and vomiting. Conversely, codeine intoxication can be anticipated with ultra-rapid CYP2D6 metabolism.

While codeine is undoubtedly not a wonder analgesic, it is still prescribed because of the belief that being a weak opioid, it is safe. There has been a recent FDA warning on codeine use in nursing mothers ( http://www.fda.gov/bbs/topics/NEWS/2007/NEW01685.html ) following the death of a breastfed 13 days old neonate thought to have suffered a morphine overdose because his mother was taking codeine and was a CYP2D6 ultra-rapid metabolizer. Since that fatal report in 2006, numerous publications have addressed the issue of potential neonatal and pediatric codeine intoxications according to pharmacokinetic phenotype. This case exemplifies very precisely the irrefutable input of diagnostic pharmacogenetic testing and carries direct clinical implications for obstetric anaesthesiologists, obstetricians and neonatologists.

Similarly, the effect of tramadol, a centrally acting μ-opioid agonist that has a dual mechanism of action, binding to the μORs and weakly inhibiting the neuronal reuptake of norepinephrine and serotonin, is also influenced by the CYP2D6 genotype and interactions with any other co-administered medication undergoing CYP2D6 metabolism should be anticipated.

Neuraxial labour analgesia and post-caesarean analgesia

The μOR, encoded by genetic locus OPRM1 , has been the focus of several genetic studies because this receptor is the primary site of action for many endogenous opioid peptides and the major target for opioid analgesics, such as morphine, heroin (diamorphine), fentanyl, and methadone. The μOR is a GPCR, and a number of SNPs have been described for OPRM1 . At nucleotide position 118, an adenine substitution by a guanine (A118 G), has been reported to occur with an allelic frequency of 10–30% among Caucasians with a higher prevalence among Asians. The major interest for this particular SNP is due to its pharmacological and physiological consequences. In vitro , the presence of at least one G118 allele has been shown to increase the binding affinity and potency of β-endorphin. Thus, individuals carrying the variant receptor gene could show differences in some of the functions mediated by β-endorphin action at the altered μOR, such as higher thresholds to pain. Consistent with this laboratory finding, one in vivo study in a human experimental pain model demonstrated that volunteers carrying a G118 allele exhibited lower sensitivity to pressure pain compared with A118 homozygotes. The exact relationship of this polymorphism with the responses to oral opioids for treatment of chronic pain, post-operative requirements of intravenous alfentanil or morphine or the adverse effects and toxicity profiles appear to go ‘in the opposite direction’. Individuals carrying the variant G118 allele may actually require higher doses of morphine, which is in contrast with what is expected if the variant μOR is truly associated with an increased endogenous opioid activity.

Our group has demonstrated that the A118 G polymorphism impacts on the median effective dose (ED50) of intrathecal fentanyl in labour; nulliparous women carrying the variant G118 allele required substantially lower doses of intrathecal fentanyl to achieve adequate analgesia early in labour (with a 1.5 to 2 fold difference between genetic groups). Of note, cervical dilatation at the time of analgesia request was significantly less in women homozygous for the wild-type variant (A118) than that in women carrying one or two variant alleles (G118). This is of interest, since women received combined-spinal epidural (CSE) analgesia by their own request, at the time they experienced painful contractions. Since, it has been demonstrated previously that epidural analgesic requirements increase with progress of labour and cervical dilatation, the expectation would be that women carrying the mutant G allele should have greater analgesic requirements due to the greater cervical dilatation at which they requested analgesia; our finding that these women require less fentanyl therefore may actually underestimate the true effect of genotype. This is consistent with the premise that women carrying the G allele may have higher pain tolerance and therefore request epidural analgesia at a later stage in labour. Our finding of a significant difference in the median effective dose (ED50) according to genotype is clinically relevant, because provision of optimal labour analgesia remains an ongoing challenge for obstetric anaesthesiologists. A major goal of labour analgesia is minimal motor impairment, emphasizing the need to minimize local anaesthetic use. Similarly, there is a need to reduce opioid doses in order to minimize opioid-related side effects such as pruritus and foetal bradycardia (which appear to be dose-dependant). Therefore, according to our findings, genotyping may well help improve the delivery of labour analgesia because a 1.5–2 fold increase in fentanyl dose is not trivial.

We have also evaluated the effect of the A118 G polymorphism on the duration of intrathecal fentanyl analgesia in early labour and did not find any difference between genotypic groups. While the A118 G polymorphism may influence drug potency, there may be no pharmacokinetic effect altering duration of drug action.

As mentioned above, our findings in labouring women are in disagreement with studies looking at post-operative intravenous consumption of morphine after general or orthopedic surgery. In addition, three studies specifically evaluated post-caesarean analgesia with intrathecal or intravenous morphine. One study did not find a difference in the duration of intrathecal morphine analgesia or need for analgesic supplementation in women carrying the G118 allele; however, among these women, pruritus was less frequent during the first 24 h post-operatively. The two other studies, both from Singapore, reported that Asian women with at least one G118 allele exhibited increased post-caesarean morphine consumption with intravenous patient-controlled analgesia.

These differences may be due to the fact that spinal and systemic opioid pharmacokinetics and pharmacodynamics may be different; enhanced analgesia in response to intrathecal fentanyl in the presence of the G118 allele may not exist in response to intravenous fentanyl or other opioids via the intravenous route. Or one could speculate that human spinal cord receptor function and signal transduction is selectively more altered by the G118 variant than supraspinal receptors. Another potential explanation and factor to bear in mind is the different nature of the nociceptive stimulus in labour versus other painful syndromes. Indeed, similar disparate results in human genetic studies of pain sensitivity have been shown to occur with other polymorphisms commonly assessed in pain studies. A polymorphism of the catechol- O -methyltransferase gene (Val158Met) has been inconsistently found to be associated with altered nociception and response to analgesic; while one study found a significant genetic association, another did not, a third described a significant effect but not for that polymorphism, and a fourth reported that findings depended on stimulus modality.

This illustrates the challenges in evaluating a genotype-phenotype association when the underlying genetic susceptibility is clearly polygenic and the phenotype is complex in itself (multifactorial, subjective, involving pain perception and/or response to analgesics).

Last, a relatively unexpected candidate gene has been suggested to modulate the perception of pain in labouring women in an ongoing study. Women who were Gly16 homozygous for the β ₂ AR reported less pain in early labour compared to others. Whether haplotypes of β ₂ AR predict more rapid and less painful labour remains to be confirmed.

Undeniably, numerous candidate genes as well as elaborate models have been suggested for the study of the genetic component of pain and several excellent reviews on the pharmacogenetics of opioids and analgesia were recently published. Nonetheless, due to the inherent complexity of studying pain (different nociceptive modalities, gender differences, limitations in extrapolating data from animal models to the response in humans, interethnic and environmental differences) in addition to the obvious polygenic nature of this, it is the design and execution of large clinical studies analyzing multiples haplotypes simultaneously that remains to be the true challenge to date. Meanwhile, genome wide association studies (GWAS) in the context of acute post-operative pain are published and researchers are actively working on gene therapies for pain. It will also be of interest to see the new insights and developments brought by more research on the SCN9A gene in the near future.