Pharmacology ConferenceAbstract IIISEX DIFFERENCES IN PHARMACOGENETICS: PHASE II ENZYMES Richard M. Weinshilboum, M.D. Almost all drugs and xenobiotics undergo biotransformation. Phase II pathways in drug and xenobiotic metabolism include conjugation reactions such as glucuronidation, sulfation, acetylation, methylation or glutathione conjugation. Virtually all phase II pathways for drug metabolism display large individual variations in activity that are due to the effects of inheritance (pharmacogenetics). In experimental animals, large sex-related differences in levels of phase II enzyme activities have been demonstrated. This presentation will compare and contrast data with regard to sex-related differences in phase II drug-metabolizing enzymes in humans and experimental animals, briefly describe our present knowledge of pharmacogenetic variations in phase II enzymes in humans and will present data -- when available -- with regard to the relationship of sex to pharmacogenetic variations in phase II enzymes in humans. Finally, possible directions for future research with regard to sex-related variation in phase II drug metabolism in humans will be discussed.
Pharmacodynamic Polymorphisms: Contribution to Variability in Drug Response? Darrell R. Abernethy, M.D., Ph.D.
Donald P. McDonnell, Ph.D. Estrogen
containing medicines have been used successfully for the past fifty
years for the treatment of conditions associated with menopause. Although
initially considered a reproductive hormone, millions of years of
clinical exposure to estrogen(s) have indicated that its influence
extends to a variety of target tissues not generally considered to
be involved in reproduction. Specifically, estrogen has positive actions
in the skeleton, the cardiovascular system and possibly the central
nervous system, activities which combine to effect a positive impact
on mortality and morbidity. However, despite the medical benefits
afforded by estrogen replacement therapy (ERT), the number of women
who initiate or remain on therapy for greater than one year is relatively
small. This is due in part to the fear that estrogens increase the
risk of getting breast cancer. Consequently, it was anticipated several
years ago that there was an unmet medical need for novel estrogen
receptor modulators which would retain the beneficial effects of estrogens
in most target organs but which were inactive in the breast. Although
the perfect tissue-selective estrogen remains to be identified progress
in this direction has been made. In the past year, for instance, we
have seen Selective Estrogen Receptor Modulators (SERMs) enter into
the clinic for the prevention of osteoporosis. Compounds of this class,
which function as estrogens in the skeletal system but oppose estrogen
action in the breast, represent the first step in developing the perfect
hormone replacement therapy medicine. In this presentation, the complex
pharmacology of SERMs and how they differ mechanistically from estradiol,
the physiological ligand of the estrogen receptor, will be discussed.
Hormones and Drug Interactions: A CNS Perspective Patricia D. Kroboth, Ph.D. The
objective of this presentation is to provide an overview of the pharmacodynamic
interactions between specific CNS active drugs and progesterone and
DHEA. In addition, a brief overview of drug interactions with estrogen
will be provided. The focus of the interactions will be on agents
that affect the GABA-receptor complex and the cholinergic system.
The importance of these interactions is only partly due to the presence
of clinical interactions. These interactions may also provide us with
insight as to how sex can affect response to drugs. Ultimately, however,
drugs serve as probes for receptors and the associated neurotransmitters
that may be involved in the pathogenesis of psychiatric or debilitating
diseases, or addictive behaviors. Sex differences in prevalence and
severity of several psychiatric diseases exist. Depression and schizophrenia
are two such diseases. Additionally, there appears to be a difference
between the sexes in difficulty with successful smoking cessation.
By briefly examining these drug interactions, intriguing, unanswered
questions will be identified regarding the role of hormones and hormone
manipulation in disease, prevention of cognitive decline, and the
potential overlay of hormones and genetic differences in affecting
the occurrence of disease or response to drugs. Michael H. Smolensky, Ph.D. The
biology of young women varies in a predictable manner over time as
utradian (high frequency and pulsatile), circadian (~24-hour), circamensual
(~monthly) and circannual (~yearly) rhythms. Numerous studies document
the role of circadian rhythms in day-night patterns of severe medical
events and the exacerbation of symptoms of chronic diseases. Circadian
rhythms also affect the response of patients to diagnostic tests.
They also are known to affect the kinetics and dynamics of therapeutic
interventions, sometimes in a profound manner, according to the time
of their ingestion, inhalation or infusion (Hrushesky et. al, Temporal
Control of Drug Delivery, NY Acad. Sci. 618, 1991; Touitou & Haus,
Biologic Rhythms in Clinical and Laboratory Medicine, Spinger-Veralg,
1992; Redfern & Lemmer, Physiology & Pharmacology of Biological
Rhythms, Springer-Verlag, 1997). Surprisingly, the study of menstrual
rhythms in the occurrence and intensity of disease has received relatively
little attention (Dalton, The premenstrual Syndrome and Progersterone
Therapy. Yearbook Medical, 1977; Case & Reid, Arch. Intern. Med.
145:1405, 1998). Indeed, the symptoms of many common medical conditions,
such respiratory (allergic rhinitis and bronchial asthma), digestive,
metabolic and neurological ones, can exhibit profound menstrual cycle
variation in their intensity. Several theories have been advanced
to explain such differences; these include predictable-in-time alteration
in endocrine status, immune function and perception. The role of menstrual
cycle-dependent differences in the kinetics and effects medications
in such patterns has yet to be explored. The dissolution characteristics
as well as the distribution, metabolism and elimination of medications
can all be affected by dosing time during the 24 hours due to influence
of the human circadian time structure. Thus, the design of studies
aimed at elucidating the role of menstrual stage on the pharmacokinetics
and dynamics of medications and other chemical agents, such as ones
found in the ambient and work environments, must be based on sound
chronobiologic and pharmacologic criteria. These former include the
standardization of subjects to a common activity-rest synchronizer
schedule and the use of marker rhythms to denote the status of menstrual
cycle stage and function, including the occurrence and quality of
ovulation. The effect of birth control interventions on physiologic
and bicohemical functions (Reinberg et. al, Chronob. Internat=l. 13:199-211,1996;
Ferin et. al. Biorhythms and Human Reproduction, Wiley, 1972) and
in turn the kinetics and effects of medications and other chemical
agents also requires investigation. Study of menstrual cycle-dependencies
of the kinetics and effects of chemical agents is complicated. It
necessitates investigative protocols that take into account the effect
of dosing time during the 24 hours as well as menstrual cycle. The
results of studies suggest the same time of day need not be representative
of the same circadian time during different stages of the menstrual
cycle (See Ferin et. al, Biorhythms and Human Reproduction, Wiley,
1972). Even the season of study could have a significant influence
on findings (See Reinberg and Smolensky, Biological Rhythms and Medicine,
Springer-Verlag, 1983). William
J. Jusko, Ph.D. The assessment of pharmacodynamic differences between men and women requires control of pharmacokinetic factors and utilization of appropriate methodology to relate response to plasma or biophase drug concentration. Some therapeutic areas have notable examples of marked sex differences in drug efficacy. For example, in the cardiovascular area, aspirin is less effective in women in prevention of stroke, perhaps related to sex-hormone-dependent differences in platelet aggregation. Tirilazad, a new agent for treatment of stroke, is also less effective in women. This was partly attributed to its more rapid clearance in women, but this factor has since been ruled out in more careful study. Sex differences in analgesic effects have been well-studied, but with diverse findings. Opioids such as pentazocine show greater efficacy for pain relief in women, but the NSAID ibuprofen exhibits better responses in men with no differences in kinetics. The immune system shows intricacies with sex differences in types of disorders, role of sex hormones, and drug responses. Women have a higher incidence of autoimmune diseases (MS, RA, SLE). Sex is sometimes a factor in drug therapy for organ transplantation with differences found in organ rejection rates and in response to drug therapy. This may be related either to differences in drug clearances (eg. corticosteroids, cyclosporine) or to intrinsic differences in lymphocyte sensitivity. Unraveling the underlying determinants of sex differences in the immune system and its responsiveness to drugs needs further study. Support:
Grant No. GM 24211 from the National Institute of General Medical
Sciences R. A. Branch, M.D., M. Romkes, Ph.D., R. Frye, Ph.D., J. Wilson, Ph.D. A wide variety of probe drugs have been advocated to evaluate in vivo activity of individual drug metabolizing enzymes. The APittsburgh Cocktail@ contribution to this field was based on the following premises: (1) Selection was made of substrates that are either entirely or predominantly metabolized by a single drug metabolizing enzyme in vitro and are suitable and safe to be given in single low doses in humans. (2) A detailed pharmacokinetic study in normal subjects was used to quantitate the fractional metabolic clearance to metabolite identified from the in vitro information. This measure was used as a gold standard of in vivo enzyme activity and was compared to a variety of single point estimates obtained from urine, blood, or both. The selection of a phenotypic trait was assessed by the correlation to fractional metabolic clearance and ease of acquisition. This phenotypic marker provides an in vivo measure of drug metabolizing activity for the enzyme of interest. (3) In order to develop a cocktail combination, it is important to ensure that simultaneous administration of multiple probes does not result in mutual drug interactions. At the present time, the Pittsburgh Cocktail consists of caffeine, mephenytoin, debrisoquine, chlorzoxazone and dapsone as probe drugs for metabolism by CYP1A2, CYP2C19, CYP2D6, CYP2E1 and CYP3A4, respectively. The use of dapsone as a probe for CYP3A4 remains controversial and is under further study as it is also metabolized by CYP2E1 and CYP2C9. We are also determining whether or not flurbiprofen can be added as a probe for CYP2C9. The cocktail study involves simultaneous drug administration, with blood samples at 0, 4 and 8 hours and an 8 hour urine collection. Due to the ease of its acquisition, it can be conducted in an outpatient or clinic setting. We are acquiring cumulative experience with this tool. We have shown that there is no evidence of mutual drug:drug interaction between the component drugs. Each measure has been remarkably stable within subjects over time in the absence of experimental perturbation, yet intersubject variance is wide. Genetic polymorphisms are confirmed for CYP2D6 and CYP2C19, but not identifiable for CYP2E1. Quantitation of mRNA in white blood cells or mucosal tissue has been shown to relate to in vivo measure of CYP2D6 and CYP2E1. The cocktail approach has proved a robust technique to demonstrate selectivity and magnitude of changes in individual enzyme activities in drug induction and drug inhibition studies. In a preliminary comparison between genders in 85 normal volunteers who have undertaken the Pittsburgh Cocktail, we have confirmed the wide intersubject variation for each phenotypic trait measure in each gender. In an initial analysis, raw observations were plotted along with 90% bootstrap confidence intervals for the ratio of median in females to median in males. These intervals were based on the Abias-corrected@ approach using 2,000 bootstrap samples. Intervals whose end points fall outside of the 0.80 - 1.25 range would not be considered evidence for bioequivalence according to Schuirmann=s Atwo one-sided tests@ approach with alpha = 0.05. Using
these Agoalposts,@ only CYP2D6 and CYP2C19 show evidence of bioequivalence.
If the criteria are relaxed to 2/3 to 3/2, then CYP2E1 and CYP1A2
would also exhibit bioequivalence between genders. However, in no
instance were the 90% confidence limits either above or below 1.0
to achieve statistical difference between genders. This analysis is
considered preliminary and merits an increase in sample size. It does,
however, question the application of rigid a priori definitions of
gender equivalence and non-equivalence in drug development. Maureen T. Cronin, Ph.D. and David Flockhart, M.D. Emerging pharmacogenetic data that describe genotype-phenotype relationships are anticipated to permit more accurate prediction of complex phenotypes from composite genotypes. These data have great potential to diminish inter-individual variability in response to medications. The next step to making a good match between a patient=s genetic profile and optimal pharmacotherapy will depend on building up and refining this nascent database by incorporating more genetic analysis and phenotype characterization into clinical trial protocols. To make this possible, molecular genetic testing must become routinely available. The ideal automated system for molecular diagnosis of pharmacogenetic traits would isolate genomic DNA from an easily accessed patient sample, carry out required PCR/labeling reactions, score polymorphisms in the PCR products, provide a composite genotype, predict phenotype and evaluate the prediction using a clinically relevant measure of therapeutic outcome. In this complex and rapidly changing environment, genotyping is best supported by hybridization-based assays on DNA microarrays. However, this method is faced with the technical challenge of providing genotyping methods capable of responding to the accelerating pace of polymorphism discovery. Evolving reliable pharmacotherapeutic predictions depends on integrating increasingly complex genotype and phenotype information and correlating it with outcomes from various therapeutic alternatives. Experience with preliminary studies designed to predict drug metabolism phenotype from CYP2D6 and CYP2C19 genotypes underscores the need for rapid array design iteration capability. New strategies have been developed to address this problem and are being demonstrated using an NAT2 model.
Computational
biology in cardiovascular medicine: insights into Thomas
J. Colatsky, Ph.D., J. Jeremy Rice, Ph.D.,
References:
Epidemiological
studies demonstrate that both the incidence and prevalence of many
mental disorders are sexually dimorphic. This holds true for changes
in cognition that accompany normal aging and neurodegenerative disease.
In both cases, Hormone Replacement Therapy improves performance of
females on verbal and spatial cognitive tasks. Thus, the decrease
in gonadal steroids that accompanies aging in females may accelerate
age related cognitive decline. |