Pharmacological Probes to Validate Biomarkers for Analgesic Drug Development.

There is an urgent need for analgesics with improved efficacy, especially in neuropathic and other chronic pain conditions. Unfortunately, in recent decades, many candidate analgesics have failed in clinical phase II or III trials despite promising preclinical results. Translational assessment tools to verify engagement of pharmacological targets and actions on compartments of the nociceptive system are missing in both rodents and humans. Through the Innovative Medicines Initiative of the European Union and EFPIA, a consortium of researchers from academia and the pharmaceutical industry was established to identify and validate a set of functional biomarkers to assess drug-induced effects on nociceptive processing at peripheral, spinal and supraspinal levels using electrophysiological and functional neuroimaging techniques. Here, we report the results of a systematic literature search for pharmacological probes that allow for validation of these biomarkers. Of 26 candidate substances, only 7 met the inclusion criteria: evidence for nociceptive system modulation, tolerability, availability in oral form for human use and absence of active metabolites. Based on pharmacokinetic characteristics, three were selected for a set of crossover studies in rodents and healthy humans. All currently available probes act on more than one compartment of the nociceptive system. Once validated, biomarkers of nociceptive signal processing, combined with a pharmacometric modelling, will enable a more rational approach to selecting dose ranges and verifying target engagement. Combined with advances in classification of chronic pain conditions, these biomarkers are expected to accelerate analgesic drug development.

IMI2-PainCare-BioPain-RCT1: study protocol for a randomized, double-blind, placebo-controlled, crossover, multi-center trial in healthy subjects to investigate the effects of lacosamide, pregabalin, and tapentadol on biomarkers of pain processing observed by peripheral nerve excitability testing (NET).

BACKGROUND: Few new drugs have been developed for chronic pain. Drug development is challenged by uncertainty about whether the drug engages the human target sufficiently to have a meaningful pharmacodynamic effect. IMI2-PainCare-BioPain-RCT1 is one of four similarly designed studies that aim to link different functional biomarkers of drug effects on the nociceptive system that could serve to accelerate the future development of analgesics. This study focusses on biomarkers derived from nerve excitability testing (NET) using threshold tracking of the peripheral nervous system. METHODS: This is a multisite single-dose, subject and assessor-blind, randomized, placebo-controlled, 4-period, 4-way crossover, pharmacodynamic (PD), and pharmacokinetic (PK) study in healthy subjects. Biomarkers derived from NET of large sensory and motor fibers and small sensory fibers using perception threshold tracking will be obtained before and three times after administration of three medications known to act on the nociceptive system (lacosamide, pregabalin, tapentadol) and placebo, given as a single oral dose with at least 1 week apart. Motor and sensory NET will be assessed on the right wrist in a non-sensitized normal condition while perception threshold tracking will be performed bilaterally on both non-sensitized and sensitized forearm skin. Cutaneous high-frequency electrical stimulation is used to induce hyperalgesia. Blood samples will be taken for pharmacokinetic purposes and pain ratings as well as predictive psychological traits will be collected. A sequentially rejective multiple testing approach will be used with overall alpha error of the primary analysis split across the two primary outcomes: strength-duration time constant (SDTC; a measure of passive membrane properties and nodal persistent Na+ conductance) of large sensory fibers and SDTC of large motor fibers comparing lacosamide and placebo. The key secondary endpoint is the SDTC measured in small sensory fibers. Remaining treatment arm effects on key NET outcomes and PK modelling are other prespecified secondary or exploratory analyses. DISCUSSION: Measurements of NET using threshold tracking protocols are sensitive to membrane potential at the site of stimulation. Sets of useful indices of axonal excitability collectively may provide insights into the mechanisms responsible for membrane polarization, ion channel function, and activity of ionic pumps during the process of impulse conduction. IMI2-PainCare-BioPain-RCT1 hypothesizes that NET can serve as biomarkers of target engagement of analgesic drugs in this compartment of the nociceptive system for future Phase 1 clinical trials. Phase 2 and 3 clinical trials could also benefit from these tools for patient stratification. TRIAL REGISTRATION: This trial was registered 25/06/2019 in EudraCT ( 2019-000942-36 ).

IMI2-PainCare-BioPain-RCT3: a randomized, double-blind, placebo-controlled, crossover, multi-center trial in healthy subjects to investigate the effects of lacosamide, pregabalin, and tapentadol on biomarkers of pain processing observed by electroencephalography (EEG).

BACKGROUND: IMI2-PainCare-BioPain-RCT3 is one of four similarly designed clinical studies aiming at profiling a set of functional biomarkers of drug effects on the nociceptive system that could serve to accelerate the future development of analgesics, by providing a quantitative understanding between drug exposure and effects of the drug on nociceptive signal processing in human volunteers. IMI2-PainCare-BioPain-RCT3 will focus on biomarkers derived from non-invasive electroencephalographic (EEG) measures of brain activity. METHODS: This is a multisite single-dose, double-blind, randomized, placebo-controlled, 4-period, 4-way crossover, pharmacodynamic (PD) and pharmacokinetic (PK) study in healthy subjects. Biomarkers derived from scalp EEG measurements (laser-evoked brain potentials [LEPs], pinprick-evoked brain potentials [PEPs], resting EEG) will be obtained before and three times after administration of three medications known to act on the nociceptive system (lacosamide, pregabalin, tapentadol) and placebo, given as a single oral dose in separate study periods. Medication effects will be assessed concurrently in a non-sensitized normal condition and a clinically relevant hyperalgesic condition (high-frequency electrical stimulation of the skin). Patient-reported outcomes will also be collected. A sequentially rejective multiple testing approach will be used with overall alpha error of the primary analysis split between LEP and PEP under tapentadol. Remaining treatment arm effects on LEP or PEP or effects on EEG are key secondary confirmatory analyses. Complex statistical analyses and PK-PD modeling are exploratory. DISCUSSION: LEPs and PEPs are brain responses related to the selective activation of thermonociceptors and mechanonociceptors. Their amplitudes are dependent on the responsiveness of these nociceptors and the state of the pathways relaying nociceptive input at the level of the spinal cord and brain. The magnitude of resting EEG oscillations is sensitive to changes in brain network function, and some modulations of oscillation magnitude can relate to perceived pain intensity, variations in vigilance, and attentional states. These oscillations can also be affected by analgesic drugs acting on the central nervous system. For these reasons, IMI2-PainCare-BioPain-RCT3 hypothesizes that EEG-derived measures can serve as biomarkers of target engagement of analgesic drugs for future Phase 1 clinical trials. Phase 2 and 3 clinical trials could also benefit from these tools for patient stratification. TRIAL REGISTRATION: This trial was registered 25/06/2019 in EudraCT ( 2019%2D%2D001204-37 ).

Acute myocarditis with normal wall motion detected with 2D speckle tracking echocardiography.

UNLABELLED: SummaryWe present the case of a 26-year-old male with acute tonsillitis who was referred for coronary angiography because of chest pain, elevated cardiac biomarkers, and biphasic T waves. The patient had no cardiovascular risk factors. Echocardiography showed no wall motion abnormalities and no pericardial effusion. 2D speckle tracking revealed distinct decreased regional peak longitudinal systolic strain in the lateral and posterior walls. Ischemic disease was extremely unlikely in view of his young age, negative family history regarding coronary artery disease, and lack of regional wall motion abnormalities on the conventional 2D echocardiogram. Coronary angiography was deferred as myocarditis was suspected. To confirm the diagnosis, cardiac magnetic resonance tomography (MRT) was performed, showing subepicardial delayed hyperenhancement in the lateral and posterior walls correlating closely with the strain pattern obtained by 2D speckle tracking echocardiography. With a working diagnosis of acute myocarditis associated with acute tonsillitis, we prescribed antibiotics and nonsteroidal anti-inflammatory drugs. The patient's clinical signs resolved along with normalization of serum creatine kinase (CK) levels, and the patient was discharged on the third day after admission. LEARNING POINTS: Acute myocarditis can mimic acute coronary syndromes.Conventional 2D echocardiography lacks specific features for detection of subtle regional wall motion abnormalities.2D speckle tracking expands the scope of echocardiography in identifying myocardial dysfunction derived from edema in acute myocarditis.

Real-time 3D echocardiographic quantification of left atrial volume: multicenter study for validation with CMR.

OBJECTIVES: We studied in a multicenter setting the accuracy and reproducibility of 3-dimensional echocardiography (3DE)-derived measurements of left atrial volume (LAV) using new, dedicated volumetric software, side by side with 2-dimensional echocardiography (2DE), using cardiac magnetic resonance (CMR) imaging as a reference. BACKGROUND: Increased LAV is associated with adverse cardiovascular outcomes. Although LAV measurements are routinely performed using 2DE, this methodology is limited because it is view dependent and relies on geometric assumptions regarding left atrial shape. Real-time 3DE is free of these limitations and accordingly is an attractive alternative for the evaluation of LAV. However, few studies have validated 3DE-derived LAV measurements against an accepted independent reference standard, such as CMR imaging. METHODS: We studied 92 patients with a wide range of LAV who underwent CMR (1.5-T) and echocardiographic imaging on the same day. Images were analyzed to obtain maximal and minimal LAV: CMR images using standard commercial tools, 2DE images using a biplane area-length technique, and 3DE images using Tomtec LA Function software. Intertechnique comparisons included linear regression and Bland-Altman analyses. Reproducibility of all 3 techniques was assessed by calculating the percentage of absolute differences in blinded repeated measurements. Kappa statistics were used to compare 2DE and 3DE classification of normal/enlarged against the CMR reference. RESULTS: 3DE-derived LAV values showed higher correlation with CMR than 2DE measurements (r = 0.93 vs. r = 0.74 for maximal LAV; r = 0.88 vs. r = 0.82 for minimal LAV). Although 2DE underestimated maximal LAV by 31 ± 25 ml and minimal LAV by 16 ± 32 ml, 3DE resulted in a minimal bias of -1 ± 14 ml for maximal LAV and 0 ± 21 ml for minimal LAV. Interobserver and intraobserver variability of 2DE and 3DE measurements of maximal LAV were similar (7% to 12%) and approximately 2 times higher than CMR (4% to 5%). 3DE classified enlarged atria more accurately than 2DE (kappa: 0.88 vs. 0.71). CONCLUSIONS: Compared with CMR reference, 3DE-derived LAV measurements are more accurate than 2DE-based analysis, resulting in fewer patients with undetected atrial enlargement.

Assessment of right ventricular function using echocardiographic speckle tracking of the tricuspid annular motion: comparison with cardiac magnetic resonance.

BACKGROUND: Assessment of right ventricular (RV) function is difficult due to the complex shape of this chamber. Tricuspid annular plane systolic excursion (TAPSE) measured with M-mode echocardiography is frequently used as an index of RV function. However, its accuracy may be limited by ultrasound beam misalignment. We hypothesized that two-dimensional (2D) speckle tracking echocardiography (STE) could provide more accurate estimates of RV function. Accordingly, STE was used to quantify tricuspid annular displacement (TAD), from which RV longitudinal shortening fraction (LSF) was calculated. These STE derived indices were compared side-by-side with M-mode TAPSE measurements against cardiac magnetic resonance (CMR) derived RV ejection fraction (EF). METHODS: Echocardiography (Philips iE33, four-chamber view) and CMR (Siemens, 1.5 T) were performed on the same day in 63 patients with a wide range of RV EF (23-70% by CMR). TAPSE was measured using M-mode echocardiography. TAD and RV LSF were obtained using STE analysis (QLAB CMQ, Philips). TAPSE, TAD and RV LSF values were compared with RV EF obtained from CMR short axis stacks. RESULTS: STE analysis required <15 seconds and was able to track tricuspid annular motion in all patients as verified visually. Correlation between RV EF and TAD (0.61 free-wall, 0.65 septal) was similar to that with M-mode TAPSE (0.63). However, STE-derived RV LSF showed a higher correlation with CMR EF (r = 0.78). CONCLUSION: RV LSF measurement by STE is fast and easy to obtain and provides more accurate evaluation of RV EF than the traditional M-mode TAPSE technique, when compared to CMR reference. (Echocardiography 2012;29:19-24).

Three-dimensional analysis of regional left ventricular endocardial curvature from cardiac magnetic resonance images.

PURPOSE: Left ventricular (LV) remodeling is usually assessed using global changes in LV volume. We hypothesized that three-dimensional analysis of regional endocardial curvature from magnetic resonance images could provide clinically useful information on localized LV remodeling. We tested this approach by investigating regional differences in endocardial curvature in normal and hypokinetic ventricles. MATERIALS AND METHODS: Images were obtained in 44 patients with normal LV function (NL, N=14), dilated cardiomyopathy (DCM, N=15) or ischemic heart disease (IHD, N=15). Local surface curvedness, normalized to take into account instantaneous LV size (C(n)), was calculated throughout the cardiac cycle and compared between segment groups: NL (N=401), IHD (N=92) and DCM (N=255). RESULTS: In all normal segments, C(n) gradually increased during systole and then decreased during diastole. While both maximum and minimum values of C(n) were comparable in the basal and midventricular segments, they were significantly higher in the four apical segments and highest in the apical cap. In addition, percent change in C(n) was higher in mid and apical compared to basal segments (P<.05). At all LV levels, C(n) values in DCM segments were lower (P<.05) than in NL and IHD segments, which were similar. In contrast, percent change in C(n) was significantly lower in both IHD and DCM segments compared to NL. CONCLUSION: Three-dimensional analysis of LV endocardial curvature yielded quantitative information on regional ventricular shape consistent with the known pathophysiology, supporting its potential clinical usefulness in the evaluation of LV remodeling.

Multimodality comparison of quantitative volumetric analysis of the right ventricle.

OBJECTIVES: We undertook volumetric analysis of the right ventricle (RV) by real-time 3-dimensional echocardiography (RT3DE), cardiac magnetic resonance (CMR), and cardiac computed tomography (CCT) on images obtained in RV-shaped phantoms and in patients with a wide range of RV geometry. BACKGROUND: Assessment of the RV by 2-dimensional (2D) echocardiography remains challenging due to its unique geometry and limitations of the current analysis techniques. RT3DE, CMR, and CCT, which can quantify RV volumes, promise to overcome the limitations of 2D echocardiography. METHODS: Images were analyzed using RV Analysis software. Volumes measured in vitro were compared with the true volumes. The human protocol included 28 patients who underwent RT3DE, CMR, and CT on the same day. Volumetric analysis of CMR images was used as a reference, against which RT3DE and CCT measurements were compared using linear regression and Bland-Altman analyses. To determine the reproducibility of the volumetric analysis, repeated measurements were performed for all 3 imaging modalities in 11 patients. RESULTS: The in vitro measurements showed that: 1) volumetric analysis of CMR images yielded the most accurate measurements; 2) CCT measurements showed slight (4%) but consistent overestimation; and 3) RT3DE measurements showed small underestimation, but considerably wider margins of error. In humans, both RT3DE and CCT measurements correlated highly with the CMR reference (r=0.79 to 0.89) and showed the same trends of underestimation and overestimation noted in vitro. All interobserver and intraobserver variability values were <14%, with those of CMR being the highest. CONCLUSIONS: Volumetric quantification of RV volume was performed on CMR, CCT, and RT3DE images. Eliminating analysis-related intermodality differences allowed fair comparisons and highlighted the unique limitations of each modality. Understanding these differences promises to aid in the functional assessment of the RV.

Quantitative evaluation of regional left ventricular function using three-dimensional speckle tracking echocardiography in patients with and without heart disease.

Although 2-dimensional (2D) speckle tracking echocardiography has been shown to be useful in the assessment of regional left ventricular function, it is limited by the assumption that speckles can be tracked frame-to-frame within the imaging plane, even though the cardiac motion is 3-dimensional (3D). Our goal was to evaluate new 3D-speckle tracking echocardiographic (STE) software by (1) comparing the regional wall motion measurements against 2D-STE images, and (2) testing its ability to identify regional wall motion abnormalities. The 2D images and real-time 3D data sets (Toshiba) obtained from 32 subjects were analyzed to measure segmental radial and longitudinal displacements and rotation, as well as the radial, longitudinal, and circumferential strains. The intertechnique comparisons included regression and Bland-Altman analyses. Additionally, cardiac magnetic resonance images (Siemens 1.5 T) acquired the same day were reviewed by an expert who classified the segments as normal or abnormal. The values of each 3D-STE index were compared between the normal and abnormal segments. The 3D-STE and 2D-STE indexes did not correlate well (r = 0.16 to 0.76) and showed wide limits in intertechnique agreement (2 SD: 5 to 6 mm for displacements, 14 degrees rotation, 17% to 52% strains) despite only minimal biases, indicating that these 2 techniques are not interchangeable. In normal segments, 3D-STE showed greater displacements, reflecting the out-of-plane motion component; smaller SDs, indicating tighter normal ranges; and a gradual decrease in radial and longitudinal displacement and a reversal in rotation from the base to the apex. In the abnormal segments, all 3D-STE indexes were reduced, reaching significance for 5 of 6 indexes. In conclusion, this is the first study to evaluate the new 3D-STE technique for measurement of regional wall motion indexes. Our findings have demonstrated its superiority over 2D-STE.

Quantification of left ventricular volumes using three-dimensional echocardiographic speckle tracking: comparison with MRI.

AIMS: Although the utility of two-dimensional (2D) speckle tracking echocardiography (STE) to quantify left ventricular (LV) volume has been demonstrated, this methodology is limited by foreshortened views, geometric modelling, and the assumption that speckles can be tracked from frame to frame, despite their out of plane motion. To circumvent these limitations, a three-dimensional (3D) speckle tracking algorithm was recently developed. Our goal was to evaluate the accuracy of the new 3D-STE side by side with 2D-STE using cardiac magnetic resonance (CMR) as a reference. METHODS AND RESULTS: Apical two- and four-chamber views (A2C and A4C) and real-time 3D datasets (Toshiba Artida 4D System) obtained in 43 patients with a wide range of LV size and function were analysed to measure LV end-systolic and end-diastolic volumes (ESV and EDV) using 2D and 3D-STE techniques. Short-axis CMR images (Siemens 1.5T scanner) acquired on the same day were analysed to obtain ESV and EDV reference values using the method of disks approximation. Reproducibility of both STE techniques was assessed using repeated measurements. While 2D-STE correlated well with CMR (r: 0.72-0.88), it underestimated LV volumes with relatively large biases (10-30 mL) and wide limits of agreement (SD: 36-51 mL), with A2C-derived measurements being worse than A4C values. The 3D-STE measurements showed higher correlation with CMR (0.87-0.92), and importantly smaller biases (1-16 mL) and narrower limits of agreement (SD: 28-37 mL). In addition, 3D-STE showed lower inter- and intra-observer variability (11-14% and 12-13%), than 2D-STE (16-17% and 12-16%, respectively). CONCLUSION: This is the first study to validate the new 3D-STE technique for LV volume measurements and demonstrate its superior accuracy and reproducibility over previously used 2D-STE technique.

Real-time 3-dimensional echocardiographic quantification of left ventricular volumes: multicenter study for validation with magnetic resonance imaging and investigation of sources of error.

OBJECTIVES: We sought to study: 1) the accuracy and reproducibility of real-time 3-dimensional echocardiographic (RT3DE) analysis of left ventricular (LV) volumes in a multicenter setting, 2) interinstitutional differences in relationship with the investigators' specific experience, and 3) potential sources of volume underestimation. BACKGROUND: Reproducibility and accuracy of RT3DE evaluation of LV volumes has not been validated in multicenter studies, and LV volumes have been reported to be underestimated compared to cardiac magnetic resonance (CMR) standard. METHODS: A total of 92 patients with a wide range of ejection fractions underwent CMR and RT3DE imaging at 4 different institutions. Images were analyzed to obtain LV end-systolic volume (ESV) and end-diastolic volume (EDV). Reproducibility was assessed using repeated analyses. The investigation of potential sources of error included: phantom imaging, intermodality analysis-related differences, and differences in LV boundary identification, such as inclusion of endocardial trabeculae and mitral valve plane in the LV volume. RESULTS: The RT3DE-derived LV volumes correlated highly with CMR values (EDV: r = 0.91; ESV: r = 0.93), but were 26% and 29% lower consistently across institutions, with the magnitude of the bias being inversely related to the level of experience. The RT3DE measurements were less reproducible (4% to 13%) than CMR measurements (4% to 7%). Minimal changes in endocardial surface position (1 mm) resulted in significant differences in measured volumes (11%). Exclusion of trabeculae and mitral valve plane from the CMR reference eliminated the intermodality bias. CONCLUSIONS: The RT3DE-derived LV volumes are underestimated in most patients because RT3DE imaging cannot differentiate between the myocardium and trabeculae. To minimize this difference, tracing the endocardium to include trabeculae in the LV cavity is recommended. With the understanding of these intermodality differences, RT3DE quantification of LV volume is a reliable tool that provides clinically useful information.

Volumetric analysis of regional left ventricular function with real-time three-dimensional echocardiography: validation by magnetic resonance and clinical utility testing.

BACKGROUND: Quantitative information on regional left ventricular volumes from real-time three-dimensional echocardiographic (RT3DE) images has significant clinical potential but needs validation. AIM: To validate these measurements against cardiac magnetic resonance (CMR) and test the feasibility of automated detection of regional wall motion (RWM) abnormalities from RT3DE data. METHODS: RT3DE (Philips) and CMR (Siemens) images were obtained from 31 patients and analysed by using prototype software to semiautomatically calculate indices of regional left ventricular function, which were compared between RT3DE and CMR (linear regression, Bland-Altman). Additionally, CMR images were reviewed by an expert, whose RWM grades were used as a reference for automated classification of segments as normal or abnormal from RT3DE and from CMR images. For each modality, normal regional ejection fraction (REF) values were obtained from 15 patients with normal wall motion. In the remaining 16 patients, REFs were compared with thresholds that were derived from patients with normal wall motion and optimised using receiver operating characteristic analysis. RESULTS: RT3DE measurements resulted in good agreement with CMR. Regional indices calculated in patients with normal wall motion varied between segments, but overall were similar between modalities. In patients with abnormal wall motion, RWM was graded as abnormal in 74% segments. CMR and RT3DE thresholds were similar (16-segment average 55 (10)% and 56 (7)%, respectively). Automated interpretation resulted in good agreement with expert interpretation, similar for CMR and RT3DE (sensitivity 0.85, 0.84; specificity 0.81, 0.78; accuracy 0.84, 0.84, respectively). CONCLUSION: Analysis of RT3DE data provides accurate quantification of regional left ventricular function and allows semiautomated detection of RWM abnormalities, which is as accurate as the same algorithm applied to CMR images.

Quantitative assessment of left ventricular size and function: side-by-side comparison of real-time three-dimensional echocardiography and computed tomography with magnetic resonance reference.

BACKGROUND: Cardiac CT (CCT) and real-time 3D echocardiography (RT3DE) are being used increasingly in clinical cardiology. CCT offers superb spatial and contrast resolution, resulting in excellent endocardial definition. RT3DE has the advantages of low cost, portability, and live 3D imaging without offline reconstruction. We sought to compare both CCT and RT3DE measurements of left ventricular size and function with the standard reference technique, cardiac MR (CMR). METHODS AND RESULTS: In 31 patients, RT3DE data sets (Philips 7500) and long-axis CMR (Siemens, 1.5 T) and CCT (Toshiba, 16-slice MDCT) images were obtained on the same day without beta-blockers. All images were analyzed to obtain end-systolic and end-diastolic volumes and ejection fractions using the same rotational analysis to eliminate possible analysis-related differences. Intertechnique agreement was tested through linear regression and Bland-Altman analyses. Repeated measurements were performed to determine intraobserver and interobserver variability. Both CCT and RT3DE measurements resulted in high correlation (r2 > 0.85) compared with CMR. However, CCT significantly overestimated end-diastolic and end-systolic volumes (26 and 19 mL; P < 0.05), resulting in a small but significant bias in ejection fraction (-2.8%). RT3DE underestimated end-diastolic and end-systolic volumes only slightly (5 and 6 mL), with no significant bias in EF (0.3%; P = 0.68). The limits of agreement with CMR were comparable for the 2 techniques. The variability in the CCT measurements was roughly half of that in either RT3DE or CMR values. CONCLUSIONS: CCT provides highly reproducible measurements of left ventricular volumes, which are significantly larger than CMR values. RT3DE measurements compared more favorably with the CMR reference, albeit with higher variability.