Toward Patient-Specific,Biologically Optimized Radiation Therapy Plans for the Treatment of Glioblastoma

Purpose

To demonstrate a method of generating patient-specific, biologically-guided radiotherapy dose plans and compare them to the standard-of-care protocol.

Methods and Materials

We integrated a patient-specific biomathematical model of glioma proliferation, invasion and radiotherapy with a multiobjective evolutionary algorithm for intensity-modulated radiation therapy optimization to construct individualized, biologically-guided plans for 11 glioblastoma patients. Patient-individualized, spherically-symmetric simulations of the standard-of-care and optimized plans were compared in terms of several biological metrics.

Results

The integrated model generated spatially non-uniform doses that, when compared to the standard-of-care protocol, resulted in a 67% to 93% decrease in equivalent uniform dose to normal tissue, while the therapeutic ratio, the ratio of tumor equivalent uniform dose to that of normal tissue, increased between 50% to 265%. Applying a novel metric of treatment response (Days Gained) to the patient-individualized simulation results predicted that the optimized plans would have a significant impact on delaying tumor progression, with increases from 21% to 105% for 9 of 11 patients.

Conclusions

Patient-individualized simulations using the combination of a biomathematical model with an optimization algorithm for radiation therapy generated biologically-guided doses that decreased normal tissue EUD and increased therapeutic ratio with the potential to improve survival outcomes for treatment of glioblastoma.     

Heart Rate Variability and Blood Pressure during Dynamic and Static Exercise at Similar Heart Rate Levels

Aim was to elucidate autonomic responses to dynamic and static (isometric) exercise of the lower limbs eliciting the same moderate heart rate (HR) response. Method: 23 males performed two kinds of voluntary exercise in a supine position at similar heart rates: static exercise (SE) of the lower limbs (static leg press) and dynamic exercise (DE) of the lower limbs (cycling). Subjective effort, systolic (SBP) and diastolic blood pressure (DBP), mean arterial pressure (MAP), rate pressure product (RPP) and the time between consecutive heart beats (RR-intervals) were measured. Time-domain (SDNN, RMSSD), frequency-domain (power in the low and high frequency band (LFP, HFP)) and geometric measures (SD1, SD2) as well as non-linear measures of regularity (approximate entropy (ApEn), sample entropy (SampEn) and correlation dimension D2) were calculated. Results: Although HR was similar during both exercise conditions (88±10 bpm), subjective effort, SBP, DBP, MAP and RPP were significantly enhanced during SE. HRV indicators representing overall variability (SDNN, SD 2) and vagal modulated variability (RMSSD, HFP, SD 1) were increased. LFP, thought to be modulated by both autonomic branches, tended to be higher during SE. ApEn and SampEn were decreased whereas D₂ was enhanced during SE. It can be concluded that autonomic control processes during SE and DE were qualitatively different despite similar heart rate levels. The differences were reflected by blood pressure and HRV indices. HRV-measures indicated a stronger vagal cardiac activity during SE, while blood pressure response indicated a stronger sympathetic efferent activity to the vessels. The elevated vagal cardiac activity during SE might be a response mechanism, compensating a possible co-activation of sympathetic cardiac efferents, as HR and LF/HF was similar and LFP tended to be higher. However, this conclusion must be drawn cautiously as there is no HRV-marker reflecting “pure” sympathetic cardiac activity.     

A High-throughput-compatible FRET-based Platform for Identification and Characterization of Botulinum Neurotoxin Light Chain Modulators

Botulinum neurotoxin (BoNT) is a potent and potentially lethal bacterial toxin that binds to host motor neurons, is internalized into the cell, and cleaves intracellular proteins that are essential for neurotransmitter release. BoNT is comprised of a heavy chain (HC), which mediates host cell binding and internalization, and a light chain (LC), which cleaves intracellular host proteins essential for acetylcholine release. While therapies that inhibit toxin binding/internalization have a small time window of administration, compounds that target intracellular LC activity have a much larger time window of administrations, particularly relevant given the extremely long half-life of the toxin. In recent years, small molecules have been heavily analyzed as potential LC inhibitors based on their increased cellular permeability relative to larger therapeutics (peptides, aptamers, etc.). Lead identification often involves high-throughput screening (HTS), where large libraries of small molecules are screened based on their ability to modulate therapeutic target function. Here we describe a FRET-based assay with a commercial BoNT/A LC substrate and recombinant LC that can be automated for HTS of potential BoNT inhibitors. Moreover, we describe a manual technique that can be used for follow-up secondary screening, or for comparing the potency of several candidate compounds.     

STEPS: A grid search methodology for optimized peptide identification filtering of MS/MS database search results

For bottom‐up proteomics, there are wide variety of database‐searching algorithms in use for matching peptide sequences to tandem MS spectra. Likewise, there are numerous strategies being employed to produce a confident list of peptide identifications from the different search algorithm outputs. Here we introduce a grid‐search approach for determining optimal database filtering criteria in shotgun proteomics data analyses that is easily adaptable to any search. Systematic Trial and Error Parameter Selection‐–referred to as STEPS‐–utilizes user‐defined parameter ranges to test a wide array of parameter combinations to arrive at an optimal “parameter set” for data filtering, thus maximizing confident identifications. The benefits of this approach in terms of numbers of true‐positive identifications are demonstrated using datasets derived from immunoaffinity‐depleted blood serum and a bacterial cell lysate, two common proteomics sample types.     

Effectiveness of Partial Sedation to Reduce Stress in Captured Mule Deer

Information garnered from the capture and handling of free-ranging animals helps advance understanding of wildlife ecology and can aid in decisions on wildlife management. Unfortunately, animals may experience increased levels of stress, injuries, and death resulting from captures (e.g., exertional myopathy, trauma). Partial sedation is a technique proposed to alleviate stress in animals during capture, yet efficacy of partial sedation for reducing stress and promoting survival post-capture remains unclear. We evaluated the effects of partial sedation on physiological, biochemical, and behavioral indicators of acute stress and probability of survival post-capture for mule deer (Odocoileus hemionus) that were captured via helicopter net-gunning in the eastern Greater Yellowstone Ecosystem, Wyoming, USA. We administered 10–30 mg of midazolam and 15 mg of azaperone intramuscularly (IM) to 32 mule deer in 2016 and 53 mule deer in 2017, and maintained a control group (captured but not sedated) of 38 mule deer in 2016 and 54 mule deer in 2017. To evaluate indicators of acute stress, we measured heart rate, blood-oxygen saturation, body temperature, respiration rate, and levels of serum cortisol. We recorded number of kicks and vocalizations of deer during handling and evaluated behavior during release. We also measured levels of fecal glucocorticoids as an indicator of baseline stress. Midazolam and azaperone did not reduce physiological, biochemical, or behavioral indicators of acute stress or influence probability of survival post-capture. Mule deer that were administered midazolam and azaperone, however, were more likely to hesitate, stumble or fall, and walk during release compared with individuals in the control group, which were more likely to trot, stot, or run without stumbling or falling. Our findings suggest that midazolam (10–30 mg IM) and azaperone (15 mg IM) may not yield physiological or demographic benefits for captured mule deer as previously assumed and may pose adverse effects that can complicate safety for captured animals, including drug-induced lethargy. Although we failed to find efficacy of midazolam and azaperone as a method for reducing stress in captured mule deer, the efficacy of midazolam and azaperone or other combinations of partial sedatives in reducing stress may depend on the dose of tranquilizer, study animal, capture setting, and how stress is defined. © 2020 The Wildlife Society.     

Evaluating δ15N–body size relationships across taxonomic levels using hierarchical models

Ecologists routinely set out to estimate the trophic position of individuals, populations, and species composing food webs, and nitrogen stable isotopes (δ¹⁵N) are a widely used proxy for trophic position. Although δ¹⁵N values are often sampled at the level of individuals, estimates and confidence intervals are frequently sought for aggregations of individuals. If individual δ¹⁵N values are correlated as an artifact of sampling design (e.g., clustering of samples in space or time) or due to intrinsic groupings (e.g., life history stages, social groups, taxonomy), such estimates may be biased and exhibit overly optimistic confidence intervals. However, these issues can be accommodated using hierarchical modeling methods. Here, we demonstrate how hierarchical models offer an additional quantitative tool for investigating δ¹⁵N variability and we explicitly evaluate how δ¹⁵N varies with body size at successively higher levels of taxonomic aggregation in a diverse fish assemblage. The models take advantage of all available data, better account for uncertainty in parameters estimates, may improve inferences on coefficients corresponding to groups with small to moderate sample sizes, and partition variation across model levels, which provides convenient summaries of the ‘importance’ of each level in terms of unexplained heterogeneity in the data. These methods can easily be applied to diet-based studies of trophic position. Although hierarchical models are well-understood and established tools, their benefits have yet to be fully reaped by stable isotope and food web ecologists. We suggest that hierarchical models can provide a robust framework for conceptualizing and statistically modeling trophic position at multiple levels of aggregation.     

Comparison of Tissue Metal Concentrations in Zucker Lean, Zucker Obese, and Zucker Diabetic Fatty Rats and the Effects of Chromium Supplementation on Tissue Metal Concentrations

Diabetes results in several metabolic changes, including alterations in the transport, distribution, excretion, and accumulation of metals. While changes have been examined in several rat models of insulin resistance and diabetes, the metal ion concentrations in the tissues of Zucker lean, Zucker obese (an insulin resistance and early stage diabetes model), and Zucker diabetic fatty (ZDF, a type 2 diabetes model) have not previously been examined in detail. The concentration of Cu, Zn, Fe, Mg, and Ca were examined in the liver, kidney, heart and spleen, and Cr concentration in the liver and kidney of these rats were examined. Zucker obese rats have a reduction in the concentration of Cu, Zn, Fe, Mg in the liver compared to ZDF and/or lean Zucker rats, presumably as a result of the increased fat content of the liver of the obese rats. ZDF rats have increased concentrations of kidney Cu compared to the lean rats, while kidney Ca concentrations are increased in the Zucker obese rats. Spleen Fe concentrations are decreased in Zucker obese rats compared to the lean rats. No effects on metal concentrations in the heart were observed between the lean, obese, and ZDF rats, and no effects on Cr concentrations were identified. Cr(III) complexes have previously been shown to have beneficial effects on the signs of insulin resistance in Zucker obese and ZDF rats. The effects of daily gavage administration of chromium picolinate ([Cr(pic)₃]) (1 mg?Cr/kg body mass), CrCl₃ (1 mg?Cr/kg body mass), and Cr3 ([Cr₃O(propionate)₆(H₂O)₃]⁺) (33 μg and 1 mg?Cr/kg body mass) on metal concentrations in these tissues were examined. Treatment with CrCl₃ and Cr3, but not [Cr(pic)₃], at 1 mg?Cr/kg resulted in a statistically significant accumulation of Cr in the kidney of lean and obese but not ZDF rats but resulted in lowering the elevated levels of kidney Cu in ZDF rats, suggesting a beneficial effect on this symptom of type 2 diabetes.     

Calmodulin-dependent activation and inactivation of anoctamin calcium-gated chloride channels

Calcium-dependent chloride channels serve critical functions in diverse biological systems. Driven by cellular calcium signals, the channels codetermine excitatory processes and promote solute transport. The anoctamin (ANO) family of membrane proteins encodes three calcium-activated chloride channels, named ANO 1 (also TMEM16A), ANO 2 (also TMEM16B), and ANO 6 (also TMEM16F). Here we examined how ANO 1 and ANO 2 interact with Ca²⁺/calmodulin using nonstationary current analysis during channel activation. We identified a putative calmodulin-binding domain in the N-terminal region of the channel proteins that is involved in channel activation. Binding studies with peptides indicated that this domain, a regulatory calmodulin-binding motif (RCBM), provides two distinct modes of interaction with Ca²⁺/calmodulin, one at submicromolar Ca²⁺ concentrations and one in the micromolar Ca²⁺ range. Functional, structural, and pharmacological data support the concept that calmodulin serves as a calcium sensor that is stably associated with the RCBM domain and regulates the activation of ANO 1 and ANO 2 channels. Moreover, the predominant splice variant of ANO 2 in the brain exhibits Ca²⁺/calmodulin-dependent inactivation, a loss of channel activity within 30 s. This property may curtail ANO 2 activity during persistent Ca²⁺ signals in neurons. Mutagenesis data indicated that the RCBM domain is also involved in ANO 2 inactivation, and that inactivation is suppressed in the retinal ANO 2 splice variant. These results advance the understanding of Ca²⁺ regulation in anoctamin Cl⁻ channels and its significance for the physiological function that anoctamin channels subserve in neurons and other cell types.     

Immune Activation and Regulation in Simian Immunodeficiency Virus-Plasmodium fragile-Coinfected Rhesus Macaques

Human immunodeficiency virus (HIV) is characterized by immune activation, while chronic malaria is associated with elevated interleukin-10 (IL-10) levels. How these apparently antagonizing forces interact in the coinfected host is poorly understood. Using a rhesus macaque model of simian immunodeficiency virus (SIV)-Plasmodium fragile coinfection, we evaluated how innate immune effector cells affect the balance between immune activation and regulation. In vitro Toll-like receptor (TLR) responses of peripheral blood myeloid dendritic cells (mDC) and monocytes were temporarily associated with acute parasitemic episodes and elevated plasma IL-10 levels. Prolonged infection resulted in a decline of mDC function. Monocytes maintained TLR responsiveness but, in addition to IL-12 and tumor necrosis factor alpha, also produced IL-10. Consistent with the role of spleen in the clearance of parasite-infected red blood cells, coinfected animals also had increased splenic IL-10 mRNA levels. The main cellular source of IL-10 in the spleens of coinfected animals, however, was not splenic macrophages but T cells, suggesting an impairment of adaptive immunity. In contrast to those in spleen, IL-10-positive cells in axillary lymph nodes of coinfected animals were predominantly mDC, reminiscent of the immunosuppressive phenotype of peripheral blood mDC. Concurrent with IL-10 induction, however, SIV infection promoted elevated systemic IL-12 levels. The continuously increasing ratio of plasma IL-12 to IL-10 suggested that the overall host response in SIV-P. fragile-coinfected animals was shifted toward immune activation versus immune regulation. Therefore, SIV-P. fragile coinfection might be characterized by earlier manifestation of immune dysfunction and exhaustion than that of single-pathogen infections. This could translate into increased morbidity in HIV-malaria-coinfected individuals.