Personalized medicines - are pharmacists ready for the challenge?

The field of personalized medicine affords multiple opportunities to pharmacists, and pharmacists have specific knowledge, skills and abilities that make them uniquely suited to advance the use of personalized medicine as a clinical tool. The pharmacy profession as a whole, however, has been slow to embrace the concept of clinical pharmacogenetics and is now facing a critical juncture that can potentially redefine the professional identity of the pharmacist. Before practice transformation can occur, however, it is important for our profession to ask and fully explore the following question: Are pharmacists ready for the challenge of personalized medicine? When assessing the readiness of pharmacy for personalized medicine, one must consider factors that are specific to the individual pharmacist as well as systematic considerations that allow pharmacists to successfully integrate personalized medicine into their individual practice area. These include factors such as education and training, competency, an attitude of engagement and adequate support and guidance. Personnel, information technology and laboratory infrastructure are also critical elements that are required, and financially sustainable practice models must be developed. Successful advancement of clinical pharmacogenetics will also require the profession to clearly define their vision of what success looks like and where it wants to be at the end of the transformational journey. Without a clear destination, we will continue to move as individuals in different directions and fail to progress as a whole. While pharmacists might not be completely ready for the challenge of pharmacogenetics, they are most certainly up to facing the challenge. The time is right and the stage is set for pharmacy to embark on another transformative journey - a journey that will redefine the role of the pharmacist and will secure a place for pharmacy in the era of personalized medicine and beyond.

The Role of the Pediatric Pharmacist in Precision Medicine and Clinical Pharmacogenomics for Children.

With the initiatives by the National Institutes of Health and the US Food and Drug Administration, pharmacogenomics is transitioning from the laboratory to patient care. Nearly 200 drug products now contain pharmacogenomic information as part of their labeling; many of these products are commonly used in the pediatric population. Because pharmacogenomic testing can provide patient-specific predictors for drug response, pharmacists are positioned to assume a leadership role in pharmacogenomic testing, clinical interpretation of results, and recommendations for individualization of drug therapy. Opportunities for pharmacists exist in both inpatient and outpatient settings, such as pharmacist-managed clinical pharmacogenomics consultation services and educating patients and families about pharmacogenomic testing. Given the potential for genetic and age-dependent factors to influence drug selection and dosing, pediatric pharmacists should be involved in the development of dosing recommendations and interprofessional practice guidelines regarding pharmacogenomic testing in pediatric patients. Opportunities to become knowledgeable and competent in pharmacogenomics extend from coursework as part of the pharmacy curriculum to postgraduate education (e.g., residencies, fellowship, continuing education). The Pediatric Pharmacy Advocacy Group acknowledges a need for increased education of both students and practicing pharmacists with consideration for infants and children.

The Role of Megalin in the Transport of Gentamicin Across BeWo Cells, an In Vitro Model of the Human Placenta.

Aminoglycosides (AG) are known to readily cross the placenta, although the mechanisms responsible for placental transport have not been characterized. Megalin is expressed in human placenta, and it is reasonable to speculate, given its role in renal AG uptake, that it is similarly involved in placental transport. However, the role of megalin in placental AG uptake has not been established. An in vitro model to study megalin-mediated placental transport has also not been previously described. The objectives of this study, therefore, were to evaluate the human choriocarcinoma (BeWo) cell line as a model to study megalin-mediated placental transport and to assess the uptake kinetics of gentamicin, an AG antibiotic, using this in vitro model. BeWo cells were grown on Transwell® plates, and megalin expression and functional activity were assessed. Uptake of (3)H-gentamicin was also evaluated in the presence and absence of megalin inhibitors. Expression of megalin protein and mRNA in BeWo cells were confirmed via immunoblot and qPCR analysis. Uptake of fluorescein isothiocyanate (FITC)-labeled bovine serum albumin (BSA) (a megalin substrate) was time-, concentration-, and temperature-dependent consistent with a transporter-mediated process. FITC-BSA uptake was also significantly reduced in the presence of unlabeled gentamicin (a megalin substrate) and sodium maleate (to induce megalin shedding) suggesting that megalin is functionally active in BeWo cells. Gentamicin uptake exhibited time and temperature dependence, saturability and Michaelis-Menten kinetics, all of which suggest a transporter-mediated process. Gentamicin uptake was also significantly reduced in the presence of the megalin inhibitors RAP and EDTA suggesting that megalin is likely involved in gentamicin uptake.

Megalin expression in human term and preterm placental villous tissues: effect of gestational age and sample processing and storage time.

INTRODUCTION: The aims of this study were to characterize megalin expression in human term and preterm placental villous tissues and to assess the impact of gestational age and sample storage on receptor expression. METHODS: Placental tissue samples were collected from pregnant women undergoing term and preterm Cesarean deliveries. Placental villous tissues were used to quantify megalin protein and mRNA expression by western blotting and quantitative polymerase chain reaction (q-PCR), respectively. Stability of megalin expression was also evaluated under various processing and storage conditions. RESULTS: Megalin mRNA was detected in term and preterm placental villous tissues. Expression in early preterm samples was 6-fold higher than in late preterm and term samples. Refrigeration of processed term samples at 4°C for up to 18h had a slight impact on megalin mRNA expression with stored samples exhibiting mRNA levels approximately 1.5-fold lower than those frozen immediately after processing. A greater decrease in mRNA expression (up to 33-fold) was observed when processed samples were snap-frozen immediately and thawed at 4°C. Processing of samples prior to refrigeration also appeared to improve mRNA stability with significantly higher expression levels noted in processed vs. unprocessed samples at all points for up to 48h. DISCUSSION: These data suggest that expression of megalin mRNA in term placental villous tissue is relatively stable for up to 18h when samples are processed immediately and refrigerated at 4°C prior to freezing. Processing prior to storage also appears to improve mRNA stability. This paper demonstrates the practical feasibility of analyzing stored tissue samples, thus, it will help with placental mRNA analysis. Additionally, megalin expression appears to vary inversely with gestational age with the greatest expression noted in the most premature samples. Age-dependent differences in placental megalin may therefore influence fetal exposure.

Challenges in the pharmacologic management of obesity and secondary dyslipidemia in children and adolescents.

The rise in childhood obesity has lead to an increased number of children with lipid abnormalities and the predominance of a combined dyslipidemic pattern characterized by a moderate-to-severe elevation in triglycerides, normal-to-mild mild elevation in LDL cholesterol and reduced HDL cholesterol. Although recently published National Heart, Lung and Blood Institute (NHLBI) guidelines represent a significant step forward in managing primary dyslipidemias in pediatric patients, there is still no general consensus regarding the pharmacologic treatment of obesity-related lipid abnormalities in children. The use of early pharmacologic intervention to control dyslipidemias and reduce cardiovascular risk in young children is only expected to increase given the steady increase in obesity and emergence of atherosclerotic disease in pre-adolescents. Despite the increasing use of lipid-lowering therapy in children over the last few years, diet and lifestyle modification remain the first line therapy. Given the challenges of instituting and maintaining lifestyle modification in pediatric patients, however, it is likely that institution of drug therapy may be required in many children. Of all the medications currently available, the fibric acid derivatives have a cholesterol lowering profile that is most likely to be effective in obese children with the high TG/low HDL phenotype and data from a recently published study of gemfibrozil in children with metabolic syndrome are promising. However, additional information regarding the short and long-term safety and efficacy of fibrate therapy in children with obesity-related lipid disorders is needed before use of these agents can be recommended.

Receptor-mediated endocytosis across human placenta: emphasis on megalin.

Receptor-mediated endocytosis plays an important role in the maternal-fetal transport of various nutrients and drugs across human placenta. Megalin, a 600 kDa endocytic receptor, is expressed in placental syncytiotrophoblasts and is thought to contribute to the transport functions of the placenta. However, the molecular mechanisms involved in megalin-mediated transplacental transport of most substrates have not yet been characterized. Megalin is most extensively expressed on the apical surface of renal proximal tubular epithelial cells, where it is involved in the reabsorption of proteins, vitamins, and polybasic drugs including aminoglycoside (AG) antibiotics. It has been suggested that megalin-mediated endocytosis is primarily responsible for accumulation of aminoglycosides (AGs) in the renal proximal tubule, which results in direct cellular injury and death. The role of megalin in the renal uptake and accumulation of aminoglycosides has therefore received much attention. It is not known, however, whether megalin is involved in the maternal-fetal transport of AGs and other commonly used polybasic drugs. Studies designed to characterize the role of megalin in transplacental transport and to understand the molecular mechanisms involved in megalin-mediated endocytosis across human placenta are therefore needed.

The role of the pediatric pharmacist in personalized medicine and clinical pharmacogenomics for children: pediatric pharmacogenomics working group.

With the initiatives by the National Institutes of Health and the Food and Drug Administration, pharmacogenomics has now moved from the laboratory to the patient bedside. Over 100 drug-products now contain pharmacogenomic information as part of their labeling. Many of these are commonly used in the pediatric population. Direct-to-consumer genetic test kits also require intervention and guidance from healthcare professionals. This increased trend towards personalized medicine mandates that healthcare professionals develop a working knowledge about pharmacogenomics and its application towards patient care. Because pharmacogenomic testing can provide patient-specific predictors for response to and safety of medications, pharmacists are positioned to play an active role in pharmacogenomic testing, clinical interpretation of results, and recommendations for individualization of drug therapy. Opportunities for pharmacists exist in both inpatient and outpatient settings, such as pharmacist-managed clinical pharmacogenomics consultation services and educating patients and families about pharmacogenomic testing. In addition to clinical roles, pharmacists may also be involved in genetically-influenced drug discovery and development. Given the potential for genetic and age-dependent factors to influence drug selection and dosing, pediatric pharmacists should be involved in the development of dosing recommendations and interprofessional practice guidelines regarding pharmacogenomic testing in pediatric patients. Opportunities to become knowledgeable and competent in pharmacogenomics span from coursework as part of the pharmacy curriculum to postgraduate education (e.g., residencies, fellowships, continuing education). However, there exists a need for additional postgraduate learning opportunities for practicing pharmacists. As a result, the Pediatric Pharmacy Advocacy Group (PPAG) acknowledges a need for increased education of both student and practicing pharmacists, with consideration of special patient populations, such as infants and children. PPAG endorses and advocates for the involvement of pediatric pharmacists in pharmacogenomic testing and in using those results to provide safe and effective medication use in pediatric patients of all ages. Additionally, PPAG strongly encourages pediatric pharmacists to take responsibility for educating patients and their families about the importance of pharmacogenomic testing and its role in the safe and effective use of medications.

The essential research curriculum for doctor of pharmacy degree programs.

In 2008, the American College of Clinical Pharmacy appointed the Task Force on Research in the Professional Curriculum to review and make recommendations on the essential research curriculum that should be part of doctor of pharmacy (Pharm.D.) degree programs. The essential research curriculum provides all students with critical and analytical thinking and lifelong learning skills, which will apply to current and future practice and stimulate some students to pursue a career in this field. Eight key curricular competencies are as follows: identifying relevant problems and gaps in pharmacotherapeutic knowledge; generating a research hypothesis; designing a study to test the hypothesis; analyzing data results using appropriate statistical tests; interpreting and applying the results of a research study to practice; effectively communicating research and clinical findings to pharmacy, medical, and basic science audiences; interpreting and effectively communicating research and clinical findings to patients and caregivers; and applying regulatory and ethical principles when conducting research or using research results. Faculty are encouraged to use research-related examples across the curriculum in nonresearch courses and to employ interactive teaching methods to promote student engagement. Examples of successful strategies used by Pharm.D. degree programs to integrate research content into the curriculum are provided. Current pharmacy school curricula allow variable amounts of time for instructional content in research, which may or may not include hands-on experiences for students to develop research-related skills. Therefore, an important opportunity exists for schools to incorporate the essential research curriculum. Despite the challenges of implementing these recommendations, the essential research curriculum will position pharmacy school graduates to understand the importance of research and its applications to practice. This perspective is provided as an aid and a challenge to those in leadership and teaching positions within schools and colleges of pharmacy.

Urine collected from diapers can be used for 2-D PAGE in infants and young children.

Urinary proteomic profiling has potential to identify candidate biomarkers of renal injury in infants provided an adequate urine sample can be obtained. Although diapers are used to obtain urine for clinical evaluation, their use for proteomic analysis has not been investigated. We therefore performed feasibility studies on the use of diaper-extracted urine for 2-D PAGE. Pediatric waste urine (2-20 mL) was applied to gel-containing, non-gel and cotton-gauze diapers and then mechanically expressed. Urine volume and total protein were measured pre- and post-extraction. Proteins were separated via 2-D PAGE following application of urine (20-40 mL) to each matrix. 2-D PAGE was also performed on clinical specimens collected using each diaper type. Differences in the adsorption and retention of urine volume and protein were noted between matrices. Non-gel and cotton-gauze diapers provided the best protein/volume recovery and the lowest interference with the Bradford assay. 2-D PAGE was also successfully completed using urine samples from both cotton fiber matrices. Conversely, samples from low-gel diapers demonstrated poor protein separation and reproducibility. Diapers containing cotton-fiber matrices appear adequate for 2-D PAGE. Qualitative and quantitative analyses of resolved proteins using replicate, high-resolution gels will be required, however, before diaper-extracted urine can be applied in proteomic profiling.

Association of the histamine N-methyltransferase C314T (Thr105Ile) polymorphism with atopic dermatitis in Caucasian children.

STUDY OBJECTIVE: To investigate potential associations between the histamine N-methyltransferase (HNMT) gene, HNMT, C314T (Thr105Ile) polymorphism and atopic dermatitis in a cohort of Caucasian children. DESIGN: Prospective, multicenter, genotype-association study. SETTING: Four academic, tertiary care medical centers within the Pediatric Pharmacology Research Unit network. PARTICIPANTS: Two hundred forty-nine Caucasian children aged 6 months-5 years with atopic dermatitis (127 patients) or without (122 control subjects). INTERVENTION: Buccal swabs (one swab/cheek) were performed to obtain epithelial cells for extraction of genomic DNA. MEASUREMENTS AND MAIN RESULTS: Data were collected on severity of atopic dermatitis, oral antihistamine treatment, and treatment response through parental report. The HNMT genotypes were successfully obtained in 116 control subjects and 122 patients with atopic dermatitis. Frequencies of the T314 variant allele (0.12 vs 0.06, p=0.04) and combined CT/TT genotype (0.24 vs 0.12, p=0.02) were significantly higher in children with atopic dermatitis compared with control subjects. Children with genotypes conferring reduced HNMT activity were 2 times more likely to have atopic dermatitis than those who were homozygous for the C314 reference allele. CONCLUSION: Increased histamine levels in patients with atopic dermatitis may result, at least in part, from reduced enzymatic inactivation via HNMT. Genetically associated reduction in histamine biotransformation may therefore contribute to the pathogenesis, persistence, and progression of atopic dermatitis. If confirmed, these data indicate that HNMT genotype might represent a common risk factor for development of atopic dermatitis, asthma, and allergic rhinitis and may be useful in identifying individuals who are candidates for early preventive pharmacotherapeutic intervention. Additional longitudinal studies will be required to assess the relationship between genotype, disease severity, and antihistamine response.

Six-month, prospective, longitudinal, open-label caffeine and dextromethorphan phenotyping study in children with growth hormone deficiency receiving recombinant human growth hormone replacement.

BACKGROUND: Recombinant human growth hormone (r-hGH) is increasingly being used in children. Although growth hormone (GH) may alter the clearance of concomitantly administered medications, its effects on individual drug-metabolizing enzymes in children have not been characterized. OBJECTIVE: The goal of this study was to assess the activities of cytochrome P450 (CYP) 1A2, N-acetyltransferase 2, xanthine oxidase, and CYP2D6 in children with isolated idiopathic GH deficiency before and 3 and 6 months after initiation of r-hGH treatment. METHODS: This 6-month, prospective, longitudinal, open-label phenotyping study was conducted at 4 academic tertiary care centers within the Pediatric Pharmacology Research Unit network. Prepubertal or early pubertal children (4-14 years) with short stature and isolated idiopathic GH deficiency were enrolled. Patients were given 4 ounces of a cola beverage and 0.5 mg/kg of dextromethorphan (DM) before and 3 and 6 months after initiation of r-hGH treatment. Urine was collected for 8 hours after probe substrate administration, and enzyme activity was assessed using validatedcaffeine/metaboliteandDM/metabolitemolar ratios. Patients with a DM/dextrorphan molar ratio > or =0.3 were classified as poor metabolizers, and those with a ratio <0.3 were classified as extensive metabolizers. Anthropometric and biochemical responses were assessed at each visit. Blood was also obtained for determination of serum insulinlike growth factor-1 (IGF-1) levels and CYP2D6 genotype. RESULTS: Fourteen patients (mean [SD] age, 11.5 [2.6] years [age range, 4.5-14.6 years]; 11 males, 3 females; 100% white; median height and weight, 131.8 cm and 29.2 kg, respectively) completed the 3 study visits. However, data from 2 patients were excluded from analysis due to procedural violations. In all patients, growth velocity and serum IGF-1 concentrations were significantly higher (P < 0.001) after r-hGH treatment (mean doses, 0.32 and 0.33 mg/kg per week at 3 and 6 months, respectively). However, molar ratio values did not significantly change after initiation of r-hGH. CONCLUSIONS: In this study population of children with isolated idiopathic GH deficiency, no significant differences in caffeine/metabolite and DM/metabolite molar ratios were observed after initiation of r-hGH treatment.

Comparison of various urine collection intervals for caffeine and dextromethorphan phenotyping in children.

Caffeine and dextromethorphan have been used successfully both alone and in combination to assess phenotype and enzyme activity in children of various ages. Previous pediatric phenotyping studies with these agents have used varying durations of urine collection. However, the minimum duration required for accurate phenotypic assessment with these compounds in children remains unknown. We calculated the cumulative metabolite recoveries and molar ratios in urine collected from children for 2, 4, 6, and 8 hours after caffeine and dextromethorphan administration to determine when respective urinary molar ratios stabilize and thus likely accurately reflect enzyme activity. Subjects (n = 24, ages 3-8 years) were given 4 oz of Coca-Cola(R) ( approximately 11.5 mg caffeine) and a single oral dose of dextromethorphan (0.5 mg/kg). Urine was collected at discrete intervals (0-2, 2-4, 4-6, and 6-8 h) during an 8-hour period, and the cumulative metabolite recoveries and urinary molar ratios were calculated. CYP2D6 genotyping was also performed in 21 of 24 subjects. In CYP2D6 extensive metabolizers, the extent of recovery for relevant metabolites was equivalent by 4 hours and represented 45% to 60% of the total amount recovered in the 8-hour period. The 2-hour CYP1A2 ratio was significantly different from those of longer collection intervals. Metabolite ratios for all other enzymes (i.e., NAT-2, XO, and CYP2D6) were independent of the duration of urine collection. These data suggest that a 4-hour urine collection is adequate for the concurrent assessment of hepatic CYP1A2, NAT-2, XO, and CYP2D6 activity in children ages 3 to 8 years who are CYP2D6 extensive metabolizers, using standard caffeine and dextromethorphan phenotyping methods. Longer collection periods may be required, however, in younger children or CYP2D6 poor metabolizers.

Activities of cytochrome P450 1A2, N-acetyltransferase 2, xanthine oxidase, and cytochrome P450 2D6 are unaltered in children with cystic fibrosis.

The activities of hepatic cytochrome P450 (CYP) 1A2, N-acetyltransferase 2 (NAT-2), xanthine oxidase (XO), and CYP2D6 were evaluated in 12 young children (aged 3-8 years) with mild cystic fibrosis (CF) and 12 age-matched healthy control subjects by use of standard caffeine and dextromethorphan phenotyping methods. Subjects were given 4 oz of Coca-Cola (approximately 35 mg caffeine) (The Coca-Cola Company, Atlanta, Ga) and a single 0.5-mg/kg dose of dextromethorphan. Urine was collected for 8 hours after biomarker administration, and enzyme activity was assessed by use of previously validated caffeine and dextromethorphan molar ratios. CYP2D6 genotyping was also performed in 10 of 12 subjects with CF and 11 of 12 control subjects. There were no significant differences in the urinary molar ratios for any of the enzyme systems evaluated. These data suggest that CF does not alter the activities of CYP1A2, NAT-2, XO, and CYP2D6. Altered biotransformation of drugs in this patient population is likely enzyme- and isoform-specific and thus is apparent for only selected compounds that are substrates for enzymes other than CYP1A2, NAT-2, XO, and CYP2D6.

Impaired recovery of hypothalamic-pituitary-adrenal axis function and hypoglycemic seizures after high-dose inhaled corticosteroid therapy in a toddler.

BACKGROUND: Corticosteroids are the treatment of choice for children with persistent reactive airway disease. In these patients, taper and discontinuation of systemic therapy is often facilitated by transition to high-dose inhaled corticosteroid treatment. OBJECTIVE: To report a case of impaired hypothalamic-pituitary-adrenal (HPA) axis recovery and adrenal crisis associated with prolonged high-dose inhaled therapy after long-term systemic corticosteroid treatment. METHODS: A 32-month-old child with severe airway obstruction and wheezing was treated with long-term daily systemic (intravenous and oral) corticosteroids followed by high-dose inhaled fluticasone (440 to 1,320 microg/day). This child presented in adrenal crisis, as evidenced by severe hypoglycemia and seizures, I day after receiving the influenza vaccine. After hydrocortisone replacement and a long taper of fluticasone, the child's adrenal function returned to normal. RESULTS: At the time of seizure, the serum glucose was 1 mg/dL. An electroencephalogram, computed tomographic scan, and magnetic resonance imaging of the brain were normal. Adrenal insufficiency was documented (morning serum cortisol, 0.6 microg/dL; after adrenocorticotrophic hormone stimulation, 8.4 microg/dL). Repeat evaluation 3 weeks after discontinuation of all corticosteroid therapy demonstrated normal HPA axis function. CONCLUSIONS: After treatment with long-term systemic steroids, high-dose inhaled corticosteroid therapy can impair recovery of the HPA axis and place patients at risk for adrenal crisis.

Current status of gene therapy for cystic fibrosis pulmonary disease.

Cystic fibrosis (CF) is a common lethal genetic disorder that affects all ethnic populations; however, it is most prevalent in Caucasians. Intensive basic research over the last 20 years has resulted in a wealth of information regarding the CF gene, its protein product and the mutational basis of disease. This increased understanding has lead to the development of gene therapy for the treatment of CF pulmonary disease. Delivery of the CF gene to the airway requires direct in vivo transfer using vectors encoding for normal CF transmembrane regulator (CFTR) protein. Several vectors are currently available for CF gene transfer and include both viral (adenoviruses, adeno-associated viruses) and non-viral (liposomal) systems. Initial clinical trials with each of these vectors have demonstrated that gene transfer to the CF airway is possible. The efficiency of transfer and duration of expression, however, have been limited. The effects of gene transfer on correction of the basic ion transport defects have also been highly variable and inconsistent, irrespective of the vector. Currently, the risk of severe immunological reactions is the primary factor limiting the clinical advancement of gene therapy. Both the adenoviral and liposomal vectors are associated with significant acute inflammatory reactions. The adenoviruses and adeno-associated viruses also elicit humoral immune responses that significantly reduce the efficiency of transgene expression and increase the risk of readministration. Several strategies are under investigation to improve the efficiency of gene transfer to the CF airway. These include overcoming local barriers in the lung, circumventing the immune response and improving vector internalization and/or uptake. Application of gene transfer in the child and possibly the fetus are also potential future clinical applications of gene therapy. However, despite considerable research with gene therapy, there is little evidence to suggest that a well tolerated and effective gene transfer method is imminent and aggressive use of conventional pharmacological therapies currently offer the greatest promise in the treatment of patients with CF.