Dan M. Roden

Estimation of the warfarin dose with clinical and pharmacogenetic data.

BACKGROUND Genetic variability among patients plays an important role in determining the dose of warfarin that should be used when oral anticoagulation is initiated, but practical methods of using genetic information have not been evaluated in a diverse and large population. We developed and used an algorithm for estimating the appropriate warfarin dose that is based on both clinical and genetic data from a broad population base. METHODS Clinical and genetic data from 4043 patients were used to create a dose algorithm that was based on clinical variables only and an algorithm in which genetic information was added to the clinical variables. In a validation cohort of 1009 subjects, we evaluated the potential clinical value of each algorithm by calculating the percentage of patients whose predicted dose of warfarin was within 20% of the actual stable therapeutic dose; we also evaluated other clinically relevant indicators. RESULTS In the validation cohort, the pharmacogenetic algorithm accurately identified larger proportions of patients who required 21 mg of warfarin or less per week and of those who required 49 mg or more per week to achieve the target international normalized ratio than did the clinical algorithm (49.4% vs. 33.3%, P<0.001, among patients requiring < or = 21 mg per week; and 24.8% vs. 7.2%, P<0.001, among those requiring > or = 49 mg per week). CONCLUSIONS The use of a pharmacogenetic algorithm for estimating the appropriate initial dose of warfarin produces recommendations that are significantly closer to the required stable therapeutic dose than those derived from a clinical algorithm or a fixed-dose approach. The greatest benefits were observed in the 46.2% of the population that required 21 mg or less of warfarin per week or 49 mg or more per week for therapeutic anticoagulation.

The drug transporter P-glycoprotein limits oral absorption and brain entry of HIV-1 protease inhibitors.

Currently available HIV-1 protease inhibitors are potent agents in the therapy of HIV-1 infection. However, limited oral absorption and variable tissue distribution, both of which are largely unexplained, complicate their use. We tested the hypothesis that P-glycoprotein is an important transporter for these agents. We studied the vectorial transport characteristics of indinavir, nelfinavir, and saquinavir in vitro using the model P-glycoprotein expressing cell lines L-MDR1 and Caco-2 cells, and in vivo after intravenous and oral administration of these agents to mice with a disrupted mdr1a gene. All three compounds were found to be transported by P-glycoprotein in vitro. After oral administration, plasma concentrations were elevated 2-5-fold in mdr1a (-/-) mice and with intravenous administration, brain concentrations were elevated 7-36-fold. These data demonstrate that P-glycoprotein limits the oral bioavailability and penetration of these agents into the brain. This raises the possibility that higher HIV-1 protease inhibitor concentrations may be obtained by targeted pharmacologic inhibition of P-glycoprotein transport activity.

Multiple Mechanisms in the Long-QT Syndrome Current Knowledge, Gaps, and Future Directions

The congenital long-QT syndrome (LQTS) is characterized by prolonged QT intervals, QT interval lability, and polymorphic ventricular tachycardia. The manifestations of the disease vary, with a high incidence of sudden death in some affected families but not in others. Mutations causing LQTS have been identified in three genes, each encoding a cardiac ion channel. In families linked to chromosome 3, mutations in SCN5A, the gene encoding the human cardiac sodium channel, cause the disease, Mutations in the human ether-à-go-go-related gene (HERG), which encodes a delayed-rectifier potassium channel, cause the disease in families linked to chromosome 7. Among affected individuals in families linked to chromosome 11, mutations have been identified in KVLQT1, a newly cloned gene that appears to encode a potassium channel. The SCN5A mutations result in defective sodium channel inactivation, whereas HERG mutations result in decreased outward potassium current. Either mutation would decrease net outward current during repolarization and would thereby account for prolonged QT intervals on the surface ECG. Preliminary data suggest that the clinical presentation in LQTS may be determined in part by the gene affected and possibly even by the specific mutation. The identification of disease genes in LQTS not only represents a major milestone in understanding the mechanisms underlying this disease but also presents new opportunities for combined research at the molecular, cellular, and clinical levels to understand issues such as adrenergic regulation of cardiac electrophysiology and mechanisms of susceptibility to arrhythmias in LQTS and other settings.

Taking the “Idio” out of “Idiosyncratic”: Predicting Torsades de Pointes

Adverse reactions to drug therapy are the hane of the practitioner, and all the more so when they occur in an idiosyncratic, or apparently unpredictable, fashion. The occurrence of torsades de pointes during treatment with action potential prolonging drugs has long fallen into this category. Progress toward understanding mechanisms underlying torsades de pointes and identifying patients at risk would go a long way to rationalizing therapy with currently available antiarrhythmic drugs and perhaps directing development of new agents.

Allelic variants in long-QT disease genes in patients with drug-associated torsades de pointes.

Background—DNA variants appearing to predispose to drug-associated “acquired” long-QT syndrome (aLQTS) have been reported in congenital long-QT disease genes. However, the incidence of these genetic risk factors has not been systematically evaluated in a large set of patients with aLQTS. We have previously identified functionally important DNA variants in genes encoding K+ channel ancillary subunits in 11% of an aLQTS cohort. Methods and Results—The coding regions of the genes encoding the pore-forming channel proteins KvLQT1, HERG, and SCN5A were screened in (1) the same aLQTS cohort (n=92) and (2) controls, drawn from patients tolerating QT-prolonging drugs (n=67) and cross sections of the Middle Tennessee (n=71) and US populations (n=90). The frequency of three common nonsynonymous coding region polymorphisms was no different between aLQTS and control subjects, as follows: 24% versus 19% for H558R (SCN5A), 3% versus 3% for R34C (SCN5A), and 14% versus 14% for K897T (HERG). Missense mutations (absent in controls) were identified in 5 of 92 patients. KvLQT1 and HERG mutations (one each) reduced K+ currents in vitro, consistent with the idea that they augment risk for aLQTS. However, three SCN5A variants did not alter INa, which argues that they played no role in the aLQTS phenotype. Conclusions—DNA variants in the coding regions of congenital long-QT disease genes predisposing to aLQTS can be identified in ≈10% to 15% of affected subjects, predominantly in genes encoding ancillary subunits.

Incidence and clinical features of the quinidine-associated long QT syndrome: Implications for patient care

Quinidine therapy is one of the most common causes of the acquired long QT syndrome and torsade de pointes. In reviewing clinical data in 24 patients with the quinidine-associated long QT syndrome, 20 of whom had torsade de pointes, we have delineated several heretofore unreported or underemphasized features. (1) This adverse drug reaction occurred either in patients who were being treated for frequent nonsustained ventricular arrhythmias or for atrial fibrillation or flutter. (2) In patients being treated for atrial fibrillation, torsade de pointes occurred only after conversion to sinus rhythm. (3) Although most patients developed the syndrome within days of starting quinidine, four had torsade de pointes during long-term quinidine therapy, usually in association with hypokalemia. (4) Because of the large experience with this entity at our institution, we have been able to estimate the risk as at least 1.5% per year. (5) Twenty of the 24 patients had at least one major, easily identifiable, associated risk factor including serum potassium below 3.5 mEq/L (four); serum potassium between 3.5 and 3.9 mEq/L (nine); high-grade atrioventricular block (four); and marked underlying, (unrecognized) QT prolongation (two). Plasma quinidine concentrations were low, being at or below the lower limit of the therapeutic range in half of patients. The ECG features typically included absence of marked QRS widening, marked QT prolongation (by definition), and a stereotypic series of cycle length changes just prior to the onset of torsade de pointes. Torsade de pointes started after the T wave of a markedly prolonged QT interval that followed a cycle that had been markedly prolonged (usually by a post ectopic pause).(ABSTRACT TRUNCATED AT 250 WORDS)

Interrelationship between substrates and inhibitors of human CYP3A and P-glycoprotein.

AbstractPurpose. CYP3A and P-gp both function to reduce the intracellular concentration of drug substrates, one by metabolism and the other by transmembrane efflux. Moreover, it has been serendipitously noted that the two proteins have many common substrates and inhibitors. In order to test this notion more fully, systematic studies were undertaken to determine the P-gp-mediated transport and inhibitory characteristics of prototypical CYP substrates. Methods. L-MDR1, LLC-PK1, and Caco-2 cells were used to evaluate established CYP substrates as potential P-gp substrates and inhibitors in vitro, and mdrla deficient mice were used to assess the in vivo relevance of P-gp-mediated transport. Results. Some (terfenadine, erythromycin and lovastatin) but not all (nifedipine and midazolam) CYP3A substrates were found to be P-gp substrates. Except for debrisoquine, none of the prototypical substrates of other common human CYP isoforms were transported by P-gp. Studies in mdrla disrupted mice confirmed that erythromycin was a P-gp substrate but the CYP3A inhibitor ketoconazole was not. In addition, CYP3A substrates and inhibitors varied widely in their ability to inhibit the P-gp-mediated transport of digoxin. Conclusions. These results indicate that the overlap in substrate specificities of CYP3A and P-gp appears to be fortuitous rather than indicative of a more fundamental relationship.

Clinical Pharmacogenetics Implementation Consortium Guidelines for CYP2C19 Genotype and Clopidogrel Therapy: 2013 Update

Cytochrome P450 (CYP)2C19 catalyzes the bioactivation of the antiplatelet prodrug clopidogrel, and CYP2C19 loss‐of‐function alleles impair formation of active metabolites, resulting in reduced platelet inhibition. In addition, CYP2C19 loss‐of‐function alleles confer increased risks for serious adverse cardiovascular (CV) events among clopidogrel‐treated patients with acute coronary syndromes (ACSs) undergoing percutaneous coronary intervention (PCI). Guideline updates include emphasis on appropriate indication for CYP2C19 genotype–directed antiplatelet therapy, refined recommendations for specific CYP2C19 alleles, and additional evidence from an expanded literature review (updates at http://www.pharmgkb.org).

Development of a Large-Scale De-Identified DNA Biobank to Enable Personalized Medicine

Our objective was to develop a DNA biobank linked to phenotypic data derived from an electronic medical record (EMR) system. An “opt‐out” model was implemented after significant review and revision. The plan included (i) development and maintenance of a de‐identified mirror image of the EMR, namely, the “synthetic derivative” (SD) and (ii) DNA extracted from discarded blood samples and linked to the SD. Surveys of patients indicated general acceptance of the concept, with only a minority (∼5%) opposing it. As a result, mechanisms to facilitate opt‐out included publicity and revision of a standard “consent to treatment” form. Algorithms for sample handling and procedures for de‐identification were developed and validated in order to ensure acceptable error rates (<0.3 and <0.1%, respectively). The rate of sample accrual is 700–900 samples/week. The advantages of this approach are the rate of sample acquisition and the diversity of phenotypes based on EMRs.

Flecainide prevents catecholaminergic polymorphic ventricular tachycardia in mice and humans.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a potentially lethal inherited arrhythmia syndrome in which drug therapy is often ineffective. We discovered that flecainide prevents arrhythmias in a mouse model of CPVT by inhibiting cardiac ryanodine receptor–mediated Ca2+ release and thereby directly targeting the underlying molecular defect. Flecainide completely prevented CPVT in two human subjects who had remained highly symptomatic on conventional drug therapy, indicating that this currently available drug is a promising mechanism-based therapy for CPVT.