Determining Effective Treatment Regimens for Breast Cancer Using Combined Immunotherapy and Chemotherapy in Vivo

Breast cancer has the highest incidence rate of all cancers globally in women, and those of African descent, especially West African females, face higher rates of triple-negative breast cancer (TNBC), a more aggressive form of breast cancer. Immunotherapy for breast cancer is a relatively new treatment option, and research is ongoing to identify the best combination treatments for increasing survival of those diagnosed with TNBC. Eganelisib (IPI-549: a PI3K-gamma inhibitor that works to shift M2 macrophages to M1 to augment T cell function) with other combinatory treatments has shown promising results in reducing tumor growth and increasing survival in mice. We have been conducting experiments to determine the most effective treatment regimen that will reduce growth of 4T1 mammary cancers, a murine TNBC model in syngeneic BalbC female mice. Combinations of eganelisib, cyclophosphamide, and anti-PD-1 or anti-PD-L1 have been tested to determine how immunotherapy and chemotherapy can induce a strong immune response, resulting in better responses to chemotherapy. Our current data indicates that a treatment regimen combining eganelisib, cyclophosphamide, and anti-PD-1 was most effective at suppressing tumor growth, compared to other treatments that only included one or two of these treatments. Mean tumor sizes of mice treated with a combination of eganelisib and other treatments were 187±70 mm3, 232±71 mm3, and 227±59 mm3 at the end of the experiments, compared to control data of 576±137 mm3, 414±31 mm3, and 591±200 mm3, respectively. These results could lead to further research on effective immunotherapy treatment combinations for TNBC.https://scholarscompass.vcu.edu/uresposters/1417/thumbnail.jp

The Impact of Torso Signal Processing on Noninvasive Electrocardiographic Imaging Reconstructions

Goal: To evaluate state-of-the-art signal processing methods for epicardial potential-based noninvasive electrocardiographic imaging reconstructions of single-site pacing data. Methods: Experimental data were obtained from two torso-tank setups in which Langendorff-perfused hearts (n = 4) were suspended and potentials recorded simultaneously from torso and epicardial surfaces. 49 different signal processing methods were applied to torso potentials, grouped as i) high-frequency noise removal (HFR) methods ii) baseline drift removal (BDR) methods and iii) combined HFR+BDR. The inverse problem was solved and reconstructed electrograms and activation maps compared to those directly recorded. Results: HFR showed no difference compared to not filtering in terms of absolute differences in reconstructed electrogram amplitudes nor median correlation in QRS waveforms (p > 0.05). However, correlation and mean absolute error of activation times and pacing site localization were improved with all methods except a notch filter. HFR applied post-reconstruction produced no differences compared to pre-reconstruction. BDR and BDR+HFR significantly improved absolute and relative difference, and correlation in electrograms (p < 0.05). While BDR+HFR combined improved activation time and pacing site detection, BDR alone produced significantly lower correlation and higher localization errors (p < 0.05). Conclusion: BDR improves reconstructed electrogram morphologies and amplitudes due to a reduction in lambda value selected for the inverse problem. The simplest method (resetting the isoelectric point) is sufficient to see these improvements. HFR does not impact electrogram accuracy, but does impact post-processing to extract features such as activation times. Removal of line noise is insufficient to see these changes. HFR should be applied post-reconstruction to ensure over-filtering does not occur

Understanding repolarization in the intracardiac unipolar electrogram: A long-lasting controversy revisited

Background: The optimal way to determine repolarization time (RT) from the intracardiac unipolar electrogram (UEG) has been a topic of debate for decades. RT is typically determined by either the Wyatt method or the “alternative method,” which both consider UEG T-wave slope, but differently.Objective: To determine the optimal method to measure RT on the UEG.Methods: Seven pig hearts surrounded by an epicardial sock with 100 electrodes were Langendorff-perfused with selective cannulation of the left anterior descending (LAD) coronary artery and submersed in a torso-shaped tank containing 256 electrodes on the torso surface. Repolarization was prolonged in the non-LAD-regions by infusing dofetilide and shortened in the LAD-region using pinacidil. RT was determined by the Wyatt (tWyatt) and alternative (tAlt) methods, in both invasive (recorded with epicardial electrodes) and in non-invasive UEGs (reconstructed with electrocardiographic imaging). tWyatt and tAlt were compared to local effective refractory period (ERP).Results: With contact mapping, mean absolute error (MAE) of tWyatt and tAlt vs. ERP were 21 ms and 71 ms, respectively. Positive T-waves typically had an earlier ERP than negative T-waves, in line with theory. tWyatt -but not tAlt-shortened by local infusion of pinacidil. Similar results were found for the non-invasive UEGs (MAE of tWyatt and tAlt vs. ERP were 30 ms and 92 ms, respectively).Conclusion: The Wyatt method is the most accurate to determine RT from (non) invasive UEGs, based on novel and historical analyses. Using it to determine RT could unify and facilitate repolarization assessment and amplify its role in cardiac electrophysiology

The circle of reentry: Characteristics of trigger-substrate interaction leading to sudden cardiac arrest

Sudden cardiac death is often caused by ventricular arrhythmias driven by reentry. Comprehensive characterization of the potential triggers and substrate in survivors of sudden cardiac arrest has provided insights into the trigger-substrate interaction leading to reentry. Previously, a “Triangle of Arrhythmogenesis”, reflecting interactions between substrate, trigger and modulating factors, has been proposed to reason about arrhythmia initiation. Here, we expand upon this concept by separating the trigger and substrate characteristics in their spatial and temporal components. This yields four key elements that are required for the initiation of reentry: local dispersion of excitability (e.g., the presence of steep repolarization time gradients), a critical relative size of the region of excitability and the region of inexcitability (e.g., a sufficiently large region with early repolarization), a trigger that originates at a time when some tissue is excitable and other tissue is inexcitable (e.g., an early premature complex), and which occurs from an excitable region (e.g., from a region with early repolarization). We discuss how these findings yield a new mechanistic framework for reasoning about reentry initiation, the “Circle of Reentry.” In a patient case of unexplained ventricular fibrillation, we then illustrate how a comprehensive clinical investigation of these trigger-substrate characteristics may help to understand the associated arrhythmia mechanism. We will also discuss how this reentry initiation concept may help to identify patients at risk, and how similar reasoning may apply to other reentrant arrhythmias

Local Translation in Primary Afferent Fibers Regulates Nociception

Recent studies have demonstrated the importance of local protein synthesis for neuronal plasticity. In particular, local mRNA translation through the mammalian target of rapamycin (mTOR) has been shown to play a key role in regulating dendrite excitability and modulating long-term synaptic plasticity associated with learning and memory. There is also increased evidence to suggest that intact adult mammalian axons have a functional requirement for local protein synthesis in vivo. Here we show that the translational machinery is present in some myelinated sensory fibers and that active mTOR-dependent pathways participate in maintaining the sensitivity of a subpopulation of fast-conducting nociceptors in vivo. Phosphorylated mTOR together with other downstream components of the translational machinery were localized to a subset of myelinated sensory fibers in rat cutaneous tissue. We then showed with electromyographic studies that the mTOR inhibitor rapamycin reduced the sensitivity of a population of myelinated nociceptors known to be important for the increased mechanical sensitivity that follows injury. Behavioural studies confirmed that local treatment with rapamycin significantly attenuated persistent pain that follows tissue injury, but not acute pain. Specifically, we found that rapamycin blunted the heightened response to mechanical stimulation that develops around a site of injury and reduced the long-term mechanical hypersensitivity that follows partial peripheral nerve damage - a widely used model of chronic pain. Our results show that the sensitivity of a subset of sensory fibers is maintained by ongoing mTOR-mediated local protein synthesis and uncover a novel target for the control of long-term pain states

Genetic Testing to Inform Epilepsy Treatment Management From an International Study of Clinical Practice

IMPORTANCE: It is currently unknown how often and in which ways a genetic diagnosis given to a patient with epilepsy is associated with clinical management and outcomes. OBJECTIVE: To evaluate how genetic diagnoses in patients with epilepsy are associated with clinical management and outcomes. DESIGN, SETTING, AND PARTICIPANTS: This was a retrospective cross-sectional study of patients referred for multigene panel testing between March 18, 2016, and August 3, 2020, with outcomes reported between May and November 2020. The study setting included a commercial genetic testing laboratory and multicenter clinical practices. Patients with epilepsy, regardless of sociodemographic features, who received a pathogenic/likely pathogenic (P/LP) variant were included in the study. Case report forms were completed by all health care professionals. EXPOSURES: Genetic test results. MAIN OUTCOMES AND MEASURES: Clinical management changes after a genetic diagnosis (ie, 1 P/LP variant in autosomal dominant and X-linked diseases; 2 P/LP variants in autosomal recessive diseases) and subsequent patient outcomes as reported by health care professionals on case report forms. RESULTS: Among 418 patients, median (IQR) age at the time of testing was 4 (1-10) years, with an age range of 0 to 52 years, and 53.8% (n = 225) were female individuals. The mean (SD) time from a genetic test order to case report form completion was 595 (368) days (range, 27-1673 days). A genetic diagnosis was associated with changes in clinical management for 208 patients (49.8%) and usually (81.7% of the time) within 3 months of receiving the result. The most common clinical management changes were the addition of a new medication (78 [21.7%]), the initiation of medication (51 [14.2%]), the referral of a patient to a specialist (48 [13.4%]), vigilance for subclinical or extraneurological disease features (46 [12.8%]), and the cessation of a medication (42 [11.7%]). Among 167 patients with follow-up clinical information available (mean [SD] time, 584 [365] days), 125 (74.9%) reported positive outcomes, 108 (64.7%) reported reduction or elimination of seizures, 37 (22.2%) had decreases in the severity of other clinical signs, and 11 (6.6%) had reduced medication adverse effects. A few patients reported worsening of outcomes, including a decline in their condition (20 [12.0%]), increased seizure frequency (6 [3.6%]), and adverse medication effects (3 [1.8%]). No clinical management changes were reported for 178 patients (42.6%). CONCLUSIONS AND RELEVANCE: Results of this cross-sectional study suggest that genetic testing of individuals with epilepsy may be materially associated with clinical decision-making and improved patient outcomes

Cardiac Electrical Dyssynchrony is Accurately Detected by Noninvasive Electrocardiographic Imaging

Poor identification of electrical dyssynchrony is postulated to be a major factor contributing to the low success rate for cardiac resynchronization therapy (CRT). To evaluate body surface mapping and electrocardiographic imaging (ECGi) to detect electrical dyssynchrony noninvasively. Langendorff-perfused pig hearts (n=11) were suspended in a human torso-shaped tank, with LBBB induced through ablation. Recordings were taken simultaneously from a 108-electrode epicardial sock and 128 electrodes embedded in the tank surface during sinus rhythm and ventricular pacing. CT provided electrode and heart positions in the tank. Epicardial unipolar electrograms were reconstructed from torso potentials using ECGi. Dyssynchrony markers from torso potentials (e.g. QRS-duration) or ECGi (total activation time (TAT), interventricular delay (D-LR) and intraventricular markers) were correlated with those recorded from the sock. LBBB was induced (n=8) and sock-derived activation maps demonstrated interventricular dyssynchrony (D-LR and TAT) in all cases (p 0.05). Although ECGi markers were significantly lower than recorded (p <0.05) there was a significant strong linear relationship between ECGi and recorded values. ECGi correctly diagnosed electrical dyssynchrony, and interventricular resynchronization in all cases. The latest site of activation was identified to 9.1±0.6 mm by ECGi. ECGi reliably and accurately detects electrical dyssynchrony, resynchronization by biventricular pacing, and the site of latest activation, providing more information than body surface potential

Investigation into the importance of using natural PVCs and pathological models for potential-based ECGI validation

Introduction: Premature ventricular contractions (PVCs) are one of the most commonly targeted pathologies for ECGI validation, often through ventricular stimulation to mimic the ectopic beat. However, it remains unclear if such stimulated beats faithfully reproduce spontaneously occurring PVCs, particularly in the case of the R-on-T phenomenon. The objective of this study was to determine the differences in ECGI accuracy when reconstructing spontaneous PVCs as compared to ventricular-stimulated beats and to explore the impact of pathophysiological perturbation on this reconstruction accuracy.Methods: Langendorff-perfused pig hearts (n = 3) were suspended in a human torso-shaped tank, and local hyperkalemia was induced through perfusion of a high-K+ solution (8 mM) into the LAD. Recordings were taken simultaneously from the heart and tank surfaces during ventricular pacing and during spontaneous PVCs (including R-on-T), both at baseline and high K+. Epicardial potentials were reconstructed from torso potentials using ECGI.Results: Spontaneously occurring PVCs were better reconstructed than stimulated beats at baseline in terms of electrogram morphology [correlation coefficient (CC) = 0.74 ± 0.05 vs. CC = 0.60 ± 0.10], potential maps (CC = 0.61 ± 0.06 vs. CC = 0.51 ± 0.12), and activation time maps (CC = 0.86 ± 0.07 vs. 0.76 ± 0.10), though there was no difference in the localization error (LE) of epicardial origin (LE = 14 ± 6 vs. 15 ± 11 mm). High K+ perfusion reduced the accuracy of ECGI reconstructions in terms of electrogram morphology (CC = 0.68 ± 0.10) and AT maps (CC = 0.70 ± 0.12 and 0.59 ± 0.23) for isolated PVCs and paced beats, respectively. LE trended worse, but the change was not significant (LE = 17 ± 9 and 20 ± 12 mm). Spontaneous PVCs were less well when the R-on-T phenomenon occurred and the activation wavefronts encountered a line of block.Conclusion: This study demonstrates the differences in ECGI accuracy between spontaneous PVCs and ventricular-paced beats. We also observed a reduction in this accuracy near regions of electrically inactive tissue. These results highlight the need for more physiologically realistic experimental models when evaluating the accuracy of ECGI methods. In particular, reconstruction accuracy needs to be further evaluated in the presence of R-on-T or isolated PVCs, particularly when encountering obstacles (functional or anatomical) which cause line of block and re-entry