Dihydropyrimidine Dehydrogenase Phenotyping Using Pretreatment Uracil: A Note of Caution Based on a Large Prospective Clinical Study.

In clinical practice, 25-30% of the patients treated with fluoropyrimidines experience severe fluoropyrimidine-related toxicity. Extensively clinically validated DPYD genotyping tests are available to identify patients at risk of severe toxicity due to decreased activity of dihydropyrimidine dehydrogenase (DPD), the rate limiting enzyme in fluoropyrimidine metabolism. In April 2020, the European Medicines Agency recommended that, as an alternative for DPYD genotype-based testing for DPD deficiency, also phenotype testing based on pretreatment plasma uracil levels is a suitable method to identify patients with DPD deficiency. Although the evidence for genotype-directed dosing of fluoropyrimidines is substantial, the level of evidence supporting plasma uracil levels to predict DPD activity in clinical practice is limited. Notwithstanding this, uracil-based phenotyping is now used in clinical practice in various countries in Europe. We aimed to determine the value of pretreatment uracil levels in predicting DPD deficiency and severe treatment-related toxicity. To this end, we determined pretreatment uracil levels in 955 patients with cancer, and assessed the correlation with DPD activity in peripheral blood mononuclear cells (PBMCs) and fluoropyrimidine-related severe toxicity. We identified substantial issues concerning the use of pretreatment uracil in clinical practice, including large between-center study differences in measured pretreatment uracil levels, most likely as a result of pre-analytical factors. Importantly, we were not able to correlate pretreatment uracil levels with DPD activity nor were uracil levels predictive of severe treatment-related toxicity. We urge that robust clinical validation should first be performed before pretreatment plasma uracil levels are used in clinical practice as part of a dosing strategy for fluoropyrimidines.

Comparative evaluation of the left ventricular mass in patients with chronic kidney disease in periodontally healthy, chronic gingivitis, and chronic periodontitis patients.

OBJECTIVES: Emerging evidence suggests that inflammation due to periodontal diseases may not be limited to adjacent oral tissues but may have influence on systemic diseases such as chronic kidney diseases (CKD) and cardiovascular diseases. Hence, this study was aimed to evaluate and compare left ventricular mass (LVM) in patients with CKD undergoing hemodialysis (CKDH) in periodontally healthy, chronic gingivitis, and chronic periodontitis. METHODOLOGY: A total of 60Â patients diagnosed with CKDH were divided equally into three groups based on periodontal status as CKDH patients with healthy periodontium (Group I), CKDH patients with chronic gingivitis (Group II), and CKDH patients with chronic periodontitis (Group III). These patients were assessed clinically, biochemically, and echocardiographically. LVM in each of these patients was calculated according to Devereux formula and was indexed to height. RESULTS: Group II and Group III patients exhibited higher mean LVM of 199.51 ± 40.17 g and 200.35 ± 65.04Â g, respectively, as compared to Group I of 161.56 ± 27.99Â g. Similarly, LVM index (LVMI) was found to be more in Group II and Group III at 59.36 ± 13.14Â g/m2.7 and 57.83 ± 19.94Â g/m2.7, respectively, while it was 45.99 ± 11.87 g/m2.7 for Group I patients. CONCLUSION: Increasing the severity of periodontal diseases in CKDH patients is associated with increase in LVM and LVMI. Periodontal screening and intervention would enable the clinician to refine cardiovascular risk assessment in such patients.

Point-of-care companion diagnostic tests for personalizing psychiatric medications: fulfilling an unmet clinical need.

Over the last decade stable isotope-labeled substrates have been used as probes for rapid, point-of-care, non-invasive and user-friendly phenotype breath tests to evaluate activity of drug metabolizing enzymes. These diagnostic breath tests can potentially be used as companion diagnostics by physicians to personalize medications, especially psychiatric drugs with narrow therapeutic windows, to monitor the progress of disease severity, medication efficacy and to study in vivo the pharmacokinetics of xenobiotics. Several genotype tests have been approved by the FDA over the last 15 years for both cytochrome P450 2D6 and 2C19 enzymes, however they have not been cleared for use in personalizing medications since they fall woefully short in identifying all non-responders to drugs, especially for the CYP450 enzymes. CYP2D6 and CYP2C19 are among the most extensively studied drug metabolizing enzymes, involved in the metabolism of approximately 30% of FDA-approved drugs in clinical use, associated with large individual differences in medication efficacy or tolerability essentially due to phenoconversion. The development and commercialization via FDA approval of the non-invasive, rapid (<60 min), in vivo, phenotype diagnostic breath tests to evaluate polymorphic CYP2D6 and CYP2C19 enzyme activity by measuring exhaled 13CO2 as a biomarker in breath will effectively resolve the currently unmet clinical need for individualized psychiatric drug therapy. Clinicians could personalize treatment options for patients based on the CYP2D6 and CYP2C19 phenotype by selecting the optimal medication at the right initial and subsequent maintenance dose for the desired clinical outcome (i.e. greatest efficacy and minimal side effects).

The effect of proton pump inhibitors on the CYP2C19 enzyme activity evaluated by the pantoprazole-13C breath test in GERD patients: clinical relevance for personalized medicine.

Patients with gastroesophageal reflux disease (GERD) are routinely prescribed one of the six FDA approved proton pump inhibitors (PPI). All of these PPI are inhibitors of CYP2C19 enzyme to varying degrees. The phenotype pantoprazole-13C breath test (Ptz-BT) was used to identify patients who are poor metabolizers (PM) and the extent of phenoconversion of CYP2C19 enzyme activity caused by four PPI (omeprazole, esomprazole pantoprazole and rabeprazole) in 54 newly diagnosed GERD patients prior to initiating randomly selected PPI therapy and 30 d after PPI therapy. The phenoconversion after 30 d of PPI therapy in GERD patients was statistically significant (p =0.001) with omeprazole/esomeprazole (n = 27) strong CYP2C19 inhibitors, while there was no change in CYP2C19 enzyme activity (p = 0.8) with pantoprazole/ rabeprazole (n = 27), weak CYP2C19 inhibitors. The concommitant use of omeprazole/esomeprazole, therefore, could have critical clinical relevance in individualizing medications metabolized primarily by CYP2C19 such as PPI, clopidogrel, phenytoin, cyclophosphamide, thalidomide, citalopram, clonazepam, diazepam, proguanil, tivantinib etc. The rapid (30 min), in vivo, and non-invasive phenotype Ptz-BT can evaluate CYP2C19 enzyme activity. More importantly, it can identify GERD patients with low CYP2C19 enzyme activity (PM), caused by PPI or other concomitant medications, who would benefit from dose adjustments to maintain efficacy and avoid toxicity. The existing CYP2C19 genotype tests cannot predict the phenotype nor can it detect phenoconversion due to non genetic factors.

Expression of periodontal inflammation into left ventricular hypertrophy in Type 2 diabetes mellitus: A cross-sectional study.

BACKGROUND: Chronic periodontitis, an inflammatory disease, is closely related to certain systemic conditions such as cardiovascular diseases, obesity, and Type 2 diabetes mellitus. These conditions, occurring as comorbidities, synergically affect periodontal tissues. AIM: This study aims to examine whether chronic gingivitis and chronic generalized severe periodontitis in patients with Type 2 diabetes mellitus are associated with increased left ventricular mass (LVM). MATERIALS AND METHODS: A total of 45 patients affected with Type 2 diabetes mellitus were recruited and divided into three groups with 15 patients each according to their periodontal status: Group I consisting of healthy individuals, Group II consisting of chronic gingivitis, and Group III consisting of chronic generalized severe periodontitis. They were assessed clinically, biochemically, and echocardiographically. LVM was calculated according to Devereux formula and was indexed to height. RESULTS: The differences in the means for LVM and LVM index (LVMI) were statistically significant in three groups with a P = 0.006 and 0.014, respectively. After adjusting for the confounders, the mean values of LVM in Group I, II, and III were 149.35 ± 35.51 g, 147.95 ± 31.59 g, and 156.36 ± 36.57 g, respectively and for LVMI, the mean values were 43.61 ± 12.16 g/m(2.7) (Group I), 47.12 ± 10.84 g/m(2.7) (Group II), and 46.34 ± 12.55 g/m(2.7) (Group III). CONCLUSIONS: A positive association between chronic generalized severe periodontitis and increased LVM in Type 2 DM patients was observed, suggesting the role of periodontal disease in the left ventricular hypertrophy.

Changes in CYP2C19 enzyme activity evaluated by the [(13)C]-pantoprazole breath test after co-administration of clopidogrel and proton pump inhibitors following percutaneous coronary intervention and correlation to platelet reactivity.

Dual antiplatelet therapy (DAPT) with clopidogrel and aspirin is used for the prevention of cardiovascular events following percutaneous coronary intervention (PCI). These agents increase the risk of gastrointestinal bleeding. To prevent these events, proton pump inhibitors (PPI) are routinely prescribed. It has been reported that with the exception of pantoprazole and dexlanzoprazole, PPIs can impede conversion of clopidogrel by cytochrome P450 2C19 (CYP2C19) to its active metabolite, a critical step required for clopidogrel efficacy. Changes in CYP2C19 enzyme activity (phenotype) and its correlation with platelet reactivity following PPI therapy has not yet been fully described. In this study we attempted to determine if the [ (13)C]-pantoprazole breath test (Ptz-BT) can evaluate changes in CYP2C19 enzyme activity (phenoconversion) following the administration of PPI in coronary artery disease (CAD) patients treated with DAPT after PCI. Thirty (30) days after successful PCI with stent placement, 59 patients enrolled in the Evaluation of the Influence of Statins and Proton Pump Inhibitors on Clopidogrel Antiplatelet Effects (SPICE) trial (ClinicalTrials.gov Identifier: NCT00930670) were recruited to participate in this sub study. Patients were randomized to one of 4 antacid therapies (omeprazole, esomeprazole. pantoprazole or ranitidine). Subjects were administered the Ptz-BT and platelet function was evaluated by vasodilator-stimulated phosphoprotein (VASP) phosphorylation and light transmittance aggregometry before and 30 d after treatment with antacid therapy. Patients randomized to esomeprazole and omeprazole had greater high on-treatment platelet reactivity and lowering of CYP2C19 enzyme activity at Day 60 after 30 d of PPI therapy. Patients randomized to ranitidine and pantoprazole did not show any changes in platelet activity or CYP 2C19 enzyme activity. In patients treated with esomeprazole and omeprazole, changes in CYP2C19 enzyme activity (phenoconversion) correlated well with changes in platelet reactivity. Co-administration of omeprazole or esomeprazole in patients treated with clopidogrel results in lower CYP2C19 enzyme activity and increased platelet reactivity as measured by VASP phosphorylation test while patients given pantoprazole or ranitidine did not show any significant changes in CYP2C19 enzyme activity and platelet reactivity.

CYP2C19 Phenoconversion by Routinely Prescribed Proton Pump Inhibitors Omeprazole and Esomeprazole: Clinical Implications for Personalized Medicine.

The phenotype pantoprazole-(13)C breath test (Ptz-BT) was used to evaluate the extent of phenoconversion of CYP2C19 enzyme activity caused by commonly prescribed proton pump inhibitors (PPI) omeprazole and esomprazole. The Ptz-BT was administered to 26 healthy volunteers and 8 stable cardiovascular patients twice at baseline and after 28 days of PPI therapy to evaluate reproducibility of the Ptz-BT and changes in CYP2C19 enzyme activity (phenoconversion) after PPI therapy. The average intrapatient interday variability in CYP2C19 phenotype (n = 31) determined by Ptz-BT was considerably low (coefficient of variation, 17%). Phenotype conversion resulted in 25 of 26 (96%) nonpoor metabolizer (non-PM) volunteers/patients as measured by the Ptz-BT at baseline and after PPI therapy. The incidence of PM status by phenotype following administration of omeprazole/esomeprazole (known inhibitors of CYP2C19) was 10-fold higher than those who are genetically PMs in the general population, which could have critical clinical implications for personalizing medications primarily metabolized by CYP2C19, such as clopidogrel, PPI, cyclophosphamide, thalidomide, citalopram, clonazepam, diazepam, phenytoin, etc. The Ptz-BT can rapidly (30 minutes) evaluate CYP2C19 phenotype and, more importantly, can identify patients with phenoconversion in CYP2C19 enzyme activity caused by nongenetic factors such as concomitant drugs.

Evaluation of CYP2D6 enzyme activity using a 13C-dextromethorphan breath test in women receiving adjuvant tamoxifen.

BACKGROUND: In tamoxifen-treated patients, breast cancer recurrence differs according to CYP2D6 genotype and endoxifen steady-state concentrations (Endx Css). The ¹³C-dextromethorphan breath test (DM-BT), labeled with ¹³C at the O-CH3 moiety, measures CYP2D6 enzyme activity. We sought to examine the ability of the DM-BT to identify known CYP2D6 genotypic poor metabolizers and examine the correlation between DM-BT and Endx Css. METHODS: DM-BT and tamoxifen pharmacokinetics were obtained at baseline, 3, and 6 months following tamoxifen initiation. Potent CYP2D6 inhibitors were prohibited. The correlation between baseline DM-BT with CYP2D6 genotype and Endx Css was determined. The association between baseline DM-BT (where values ≤0.9 is an indicator of poor in vivo CYP2D6 metabolism) and Endx Css (using values≤11.2 known to be associated with poorer recurrence free survival) was explored. RESULTS: A total of 91 patients were enrolled and 77 were eligible. CYP2D6 genotype was positively correlated with baseline, 3, and 6 months DM-BT (r ranging from 0.457-0. 60; P<0.001). Both CYP2D6 genotype (r=0.47, 0.56, P<0.0001), and baseline DM-BT (r=0.60, 0.54, P<0.001) were associated with 3 and 6 months Endx Css, respectively. Seven (78%) of nine patients with low (≤11.2 nmol/l) 3 month Endx Css also had low DM-BT (≤0.9) including 2/2 CYP2D6 PM/PM and 5/5 IM/PM. In contrast, one (2%) of 48 patients with a low DM-BT had Endx Css more than 11.2 nmol/l. CONCLUSION: In patients not taking potent CYP2D6 inhibitors, DM-BT was associated with CYP2D6 genotype and 3 and 6 months Endx Css but did not provide better discrimination of Endx Css compared with CYP2D6 genotype alone. Further studies are needed to identify additional factors which alter Endx Css.

Breath tests to phenotype drug disposition in oncology.

Breath tests (BTs) have been investigated as diagnostic tools to phenotype drug disposition in cancer patients in the pursuit to individualize drug treatment. The choice of the right phenotype probe is crucial and depends on the metabolic pathway of the anticancer agent of interest. BTs using orally or intravenously administered selective non-radioactive (13)C-labeled probes to non-invasively evaluate dihydropyrimidine dehydrogenase, cytochrome P450 (CYP) 3A4, and CYP2D6 enzyme activity have been published. Clinically, a (13)C-dextromethorphan BT to predict endoxifen levels in breast cancer patients and a (13)C-uracil BT to predict fluoropyrimidine toxicity in colorectal cancer patients are most promising. However, the clinical benefit and cost effectiveness of these phenotype BTs need to be determined in order to make the transition from an experimental setting to clinical practice as companion diagnostic tests.

Regulatory issues on breath tests and updates of recent advances on [13C]-breath tests.

Over the last decade non invasive diagnostic phenotype [(13)C]-breath tests as well as tests using endogenous volatile organic compounds (VOCs) in breath have been researched extensively. However, only three breath tests have been approved by the FDA over the last 15 years. Despite the potential benefits of these companion diagnostic tests (CDx) for evaluation of drug metabolizing enzyme activities and standalone diagnostic tests for disease diagnosis to personalize medicine, the clinical and commercial development of breath tests will need to overcome a number of regulatory, financial and scientific hurdles prior to their acceptance into routine clinical practice. The regulatory agencies (FDA and EMEA) need to adapt and harmonize their approval process for companion diagnostic tests as well as standalone diagnostic breath tests for personalized medicine. The Center for Devices and Radiological Health has deemed any breath test that involves a labeled (13)C substrate/drug and a device requires a Pre Market Approval (PMA), which is analogous to an approved New Drug Application. A PMA is in effect, a private license granted to the applicant for marketing a particular medical device. Any breath test with endogenous VOCs along with a device can be approved via the 510(k) application. A number of (13)C breath tests with clinical applications have been researched recently and results have been published in reputed journals. Diagnostic companies will need to invest the necessary financial resources to develop and get regulatory approval for diagnostic breath tests capable of identifying responders/non responders for FDA approved drugs with narrow therapeutic indices (personalized medicine) or for evaluating the activity of drug metabolizing P450 polymorphic enzymes or for diagnosing diseases at an early stage or for monitoring the efficacy of medications. The financial success of these diagnostic breath tests will then depend entirely on how the test is marketed to physicians, healthcare organizations, payers (reimbursement), insurance companies and most importantly to patients, the eventual beneficiaries.

Is (+)-[13C]-pantoprazole better than (±)-[13C]-pantoprazole for the breath test to evaluate CYP2C19 enzyme activity?

Recently, we have shown that the (+)-[(13)C]-pantoprazole is more dependent on CYP2C19 metabolic status than (-)-[(13)C]-pantoprazole. In this study, we tested the hypothesis that (+)-[(13)C]-pantoprazole is a more sensitive and selective probe for evaluating CYP2C19 enzyme activity than the racemic mixture. (+)-[(13)C]-pantoprazole (95 mg) was administered orally in a sodium bicarbonate solution to healthy volunteers. Breath and plasma samples were collected before and up to 720 min after dosing. The (13)CO2 in exhaled breath samples was measured by infrared spectrometry. Ratios of (13)CO2/(12)CO2 after (+)-[(13)C]-pantoprazole relative to (13)CO2/(12)CO2 at baseline were expressed as delta over baseline (DOB). (+)-[(13)C]-pantoprazole concentrations were measured by HPLC. Genomic DNA extracted from whole blood was genotyped for CYP2C19*2, *3 and *17 using Taqman assays. Statistically significant differences in the area under the plasma concentration time curve (AUCplasma(0-∞) (p < 0.001) and oral clearance (<0.01) of (+)-[(13)C]-pantoprazole as well as in the breath test indices (delta over baseline, DOB30; and area under the DOB versus time curve, AUCDOB(0-120)) (p < 0.01) were observed among poor, intermediate and extensive metabolizer of CYP2C19. DOB30 and AUCDOB(0-120) adequately distinguished poor metabolizer from intermediate and extensive metabolizer of CYP2C19. Breath test indices significantly correlated with plasma elimination parameters of (+)-[(13)C]-pantoprazole (Pearson correlations: -0.68 to -0.73). Although relatively higher breath test indices were observed after administration of (+)-[(13)C]-pantoprazole (this study) than after (±)-[(13)C]-pantoprazole (previous study), the performance of the racemic and the enantiomer as marker of CYP2C19 activity remained similar. Our data confirm that the metabolism of (+)-[(13)C]-pantoprazole is highly dependent on CYP2C19 metabolic status, but the breath test derived from it is not superior to the racemic [(13)C]-pantoprazole in evaluating CYP2C19 activity in vivo. Thus, racemic [(13)C]-pantoprazole which is relatively easy to synthesize and more stable than (+)-[(13)C]-pantoprazole is adequate as a probe of this enzyme.

A rapid non invasive L-DOPA-¹³C breath test for optimally suppressing extracerebral AADC enzyme activity - toward individualizing carbidopa therapy in Parkinson's disease.

BACKGROUND: Peripheral carbidopa (CD) levels directly impact on central dopamine (DA) production in Parkinson disease (PD) through extracerebral inhibition of dopa decarboxylase (AADC) resulting in an increase in levodopa (LD) bioavailability. OBJECTIVE: Recent data suggests that higher CD doses than those presently used in PD treatment may result in improved clinical response. Optimizing CD doses in individual patients may, therefore, result in ideal individualized treatment. METHODS: A single center, randomized, double-blind study was carried out recruiting 5 Parkinson's disease (PD) patients already on LD/CD and 1 treatment näve PD patient using stable isotope labeled LD-1-¹³C as a substrate for a noninvasive breath test to evaluate individual AADC enzyme activity. Each patient was studied five times, receiving 200 mg LD-¹³C at each visit along with one of five randomized CD doses (0, 25, 50, 100 and 200 mg). The metabolite ¹³CO2 in breath was measured for evaluating AADC enzyme activity and plasma metabolite levels for LD-¹³C and homovanillic acid (HVA) were measured for 4 hours. RESULTS: HVA in plasma and ¹³CO2 in breath are metabolic products of LD. We found a significant positive correlation of ¹³CO2 DOB AUC0-240 with serum HVA AUC0-240 following the oral dose of LD-1-¹³C for all 5 doses of CD (r² = 0.9378). With increasing inhibition of AADC enzyme activity with CD, we observed an increase in the plasma concentration of LD.We found an inverse correlation of the 13CO2 DOB AUC with serum LD-¹³C AUC. Our studies indicate the optimal dose of CD for maximal suppression of AADC enzyme activity can be determined for each individual from ¹³CO2 generation in breath. CONCLUSIONS: The LD-breath test can be a useful noninvasive diagnostic tool for evaluation of AADC enzyme activity using the biomarker ¹³CO2 in breath, a first step in personalizing CD doses for PD patients.

Stereoselective pharmacokinetics of stable isotope (+/-)-[13C]-pantoprazole: Implications for a rapid screening phenotype test of CYP2C19 activity.

AIMS: We have previously shown that the (±)-[(13) C]-pantoprazole breath test is a promising noninvasive probe of CYP2C19 activity. As part of that trial, plasma, breath test indices and CYP2C19 (*2, *3, and *17) genotype were collected. Here, we examined whether [(13) C]-pantoprazole exhibits enantioselective pharmacokinetics and whether this enantioselectivity is correlated with indices of breath test. METHODS: Plasma (-)- and (+)-[(13) C]-pantoprazole that were measured using a chiral HPLC were compared between CYP2C19 genotypes and correlated with breath test indices. RESULTS: The AUC( 0-∞) of (+)-[(13) C]-pantoprazole in PM (*2/*2, n = 4) was 10.1- and 5.6-fold higher that EM (*1/*1or *17, n = 10) and IM (*1/*2or *3, n = 10) of CYP2C19, respectively (P < 0.001). The AUC( 0-∞) of (-)-[(13) C]-pantoprazole only significantly differed between PMs and EMs (1.98-fold; P = 0.05). The AUC( 0-∞) ratio of (+)-/(-)-[(13) C]-pantoprazole was 3.45, 0.77, and 0.67 in PM, IM, and EM genotypes, respectively. Breath test index, delta over baseline show significant correlation with AUC( 0-∞) of (+)-[(13) C]-pantoprazole (Pearson's r = 0.62; P < 0.001). CONCLUSIONS: [(13) C]-pantoprazole exhibits enantioselective elimination. (+)-[(13) C]-pantoprazole is more dependent on CYP2C19 metabolic status and may serve as a more attractive probe of CYP2C19 activity than (-)-[(13) C]-pantoprazole or the racemic mixture.

Breath biomarkers for personalized medicine.

Personalized medicine, in the near future, has the potential to revolutionize healthcare by allowing physicians to individualize therapy for patients through the early diagnosis of disease and risk assessment to optimize clinical response with minimal toxicity. The identification of biomarkers could detect, diagnose and help guide therapy to improve survival and quality of life by the early identification of responders to the drugs. Volatile organic compounds and stable isotope-labeled 13CO2 in breath can be uniquely utilized as in vivo diagnostic biomarkers of disease and/or lack of enzyme activity to aid physicians to personalize medication. Noninvasive detection of ailments and monitoring therapy by human breath analysis is an emerging field of medical diagnostics representing a rapid, economic and simple alternative to standard invasive blood analysis, endoscopy or harmful imaging techniques such as x-ray and CT scans.

Single time point diagnostic breath tests: a review.

The metabolism of ingested xenobiotics is clinically significant to minimize risk and optimize therapeutic benefits. A majority of the drugs approved by the FDA are metabolized by phase I enzymes. Stable isotope-labeled xenobiotics can be used to provide rapid in vivo phenotype assessment of phase I enzymes. In this paper, we describe three simple, noninvasive phenotype breath tests using [13C]-dextromethorphan and [13C]-pantoprazole for assessing polymorphic CYP2D6 and CYP2C19 enzyme activity and [13C]-uracil to assess the enzyme activity of DPD, DPHD and ß-ureidopropionase for identifying pyrimidine metabolic disorder. The results of the [13C]-dextromethorphan, [13C]-pantoprazole and [13C]-uracil breath test studies suggest that they have great potential for evaluating CYP2D6, CYP2C19 and DPD enzyme activities in a relatively short time with a single time point breath collection in a clinic or hospital setting. This would enable physicians to prescribe personalized therapy for each individual by selecting the ideal medication at the right dose for optimal efficacy of xenobiotics metabolized by these enzymes.

(13)C breath tests in personalized medicine: fiction or reality?

The concept of personalized medicine is gathering momentum as various biomarkers are being discovered and developed to lead to genotype and phenotype diagnostic tests, which will enable physicians to individualize therapy. Noninvasive, rapid (13)C breath tests have the potential to serve as clinically significant diagnostic tools, especially for evaluating the enzyme activity of polymorphic enzymes. This would enable physicians to rapidly identify responders/nonresponders to various drugs primarily metabolized by these enzymes prior to initiation of therapy. With the information on enzyme activity, the physician can prescribe the right drug, at the right dose, at the right time, to the right individual, for the right clinical outcome. However, the promise of the era of personalized medicine, including the novel (13)C breath tests, will have to overcome several regulatory, business and financial hurdles for diagnostic tests to become part of routine mainstream clinical practice over the next decade.

Rapid identification of the hepatic cytochrome P450 2C19 activity using a novel and noninvasive [13C]pantoprazole breath test.

We tested the hypothesis that the stable isotope [(13)C]pantoprazole is O-demethylated by cytochrome P450 CYP2C19 and that the (13)CO(2) produced and exhaled in breath as a result can serve as a safe, rapid, and noninvasive phenotyping marker of CYP2C19 activity in vivo. Healthy volunteers who had been genotyped for the CYP2C19(*)2, CYP2C19(*)3, and CYP2C19(*)17 alleles were administered a single oral dose of [(13)C]pantoprazole sodium-sesquihydrate (100 mg) with 2.1 g of sodium bicarbonate. Exhaled (13)CO(2) and (12)CO(2) were measured by IR spectroscopy before (baseline) and 2.5 to 120 min after dosing. Ratios of (13)CO(2)/(12)CO(2) after [(13)C]pantoprazole relative to (13)CO(2)/(12)CO(2) at baseline were expressed as change over baseline (DOB). Maximal DOB, DOB(15) to DOB(120), and area under the DOB versus time curve (AUC(0-120) and AUC(0-infinity)) were significantly different among three genotype groups (CYP2C19(*)1/(*)1, n = 10; CYP2C19(*)1/(*)2 or CYP2C19(*)1/(*)3, n = 10; and CYP2C19(*)2/(*)2, n = 5) with predicted extensive metabolizers (EMs), intermediate metabolizers (IMs), and poor metabolizers (PMs) of CYP2C19, respectively (Kruskal-Wallis test, p < 0.01); linear regression analysis indicated a gene-dose effect relationship (r(2) ranged between 0.236 and 0.522; all p < 0.05). These breath test indices were significantly lower in PMs than IMs (p < 0.05) or EMs (p < 0.01) of CYP2C19. [(13)C]Pantoprazole plasma exposure showed significant inverse correlation with breath test indices in the respective subjects (Pearson r = -0.74; p = 0.038). These feasibility data suggest that the [(13)C]pantoprazole breath test is a reliable, rapid, and noninvasive probe of CYP2C19 and seems to be a useful tool to optimize drug therapy metabolized by CYP2C19.

Breath tests with (13)C substrates.

Labeled (stable and radio) isotope compounds ((2)H, (3)H, (14)C, (13)C, (15)N) have been widely used as diagnostic probes in research laboratories for over 30 years in the fields of gastroenterology, hepatology, oncology, and nutrition, as well as pharmacokinetic studies in the development of drugs. (13)C stable isotope diagnostic probes are now being expanded in their scope, to provide precise evaluations of the presence or absence of etiologically significant changes in metabolism due to a specific disease or the lack of a specific enzyme. The concept exploits the use of the (13)C- label that is incorporated at the appropriate site into a selected substrate specifically designed for the targeted enzyme in a discrete metabolic pathway. The enzyme-substrate interaction results in the release of (13)CO(2) in the expired breath. The subsequent quantification of (13)CO(2) allows for the indirect determination of pharmacokinetics and the evaluation of enzyme activity. Breath tests, although non-invasive, have not been integrated routinely in clinical practice due to most of them requiring multiple breath sample collection over an extended time period. The use of area-under-the-curve (AUC) and percent-dose-recovery (PDR) parameters of breath tests to differentiate between controls and patients has been a huge barrier to implementing them into routine clinical practice due to time constraints on clinical staff. In order to get breath tests accepted in clinical practice as in vivo diagnostic tools, the tests need to be accurate with high sensitivity and specificity with a single time point breath collection post ingestion of a (13)C substrate. It is now incumbent on diagnostic test companies to invest capital for the development of promising single time point breath tests and getting regulatory board approval (FDA, EMEA), CPT codes and reimbursement. Following regulatory approvals, the breath tests would also need to be marketed aggressively by making physicians, patients, and insurance companies aware of the medical benefits to patients and lowering of healthcare costs. The diagnostic breath tests will enable physicians and patients to benefit from rapid, novel and non-invasive ways to detect enzyme deficiencies, to monitor the progress of disease severity or medication efficacy, to trace acquired and/or congenital metabolic defects, to study in vivo the pharmacokinetics of xenobiotics, and to optimize individually tailored treatment regimens for drugs with narrow therapeutic windows. The primary reason for publishing this special section on (13)C breath tests is to highlight some of the recent advances in the field of breath tests as well as to review the literature.