Pneumonitis After Concurrent Chemoradiation and Immune Checkpoint Inhibition in Patients with Locally Advanced Non-small Cell Lung Cancer.

AIMS: Pneumonitis is a common and potentially deadly complication of combined chemoradiation and immune checkpoint inhibition (CRT-ICI) in patients with locally advanced non-small cell lung cancer (LA-NSCLC). In this study we sought to identify the risk factors for pneumonitis with CRT-ICI therapy in LA-NSCLC cases and determine its impact on survival. MATERIALS AND METHODS: We conducted a retrospective chart review of 140 patients with LA-NSCLC who underwent curative-intent CRT-ICI with durvalumab between 2018 and 2021. Pneumonitis was diagnosed by a multidisciplinary team of clinical experts. We used multivariable cause-specific hazard models to identify risk factors associated with grade ≥2 pneumonitis. We constructed multivariable Cox proportional hazard models to investigate the impact of pneumonitis on all-cause mortality. RESULTS: The median age of the cohort was 67 years; most patients were current or former smokers (86%). The cumulative incidence of grade ≥2 pneumonitis was 23%. Among survivors, 25/28 patients had persistent parenchymal scarring. In multivariable analyses, the mean lung dose (hazard ratio 1.14 per Gy, 95% confidence interval 1.03-1.25) and interstitial lung disease (hazard ratio 3.8, 95% confidence interval 1.3-11.0) increased the risk for pneumonitis. In adjusted models, grade ≥2 pneumonitis (hazard ratio 2.5, 95% confidence interval 1.0-6.2, P = 0.049) and high-grade (≥3) pneumonitis (hazard ratio 8.3, 95% confidence interval 3.0-23.0, P < 0.001) were associated with higher all-cause mortality. CONCLUSIONS: Risk factors for pneumonitis in LA-NSCLC patients undergoing CRT-ICI include the mean radiation dose to the lung and pre-treatment interstitial lung disease. Although most cases are not fatal, pneumonitis in this setting is associated with markedly increased mortality.

Elimination of Ehrlich tumours by ATP-induced growth inhibition, glutathione depletion and X-rays.

ATP-induced tumour growth inhibition is accompanied by a selective decrease in the content of the tripeptide glutathione (GSH) within the cancer cells in vivo. Depletion of cellular GSH sensitizes tumours to chemotherapy and radiation, but the usefulness of this depletion depends on whether the levels of GSH can be reduced in the tumour relative to normal tissues. We report here that administration of ATP in combination with diethylmaleate and X-rays leads to complete regression of 95% of Ehrlich ascites tumours in mice. This shows that an aggressive tumour can be eliminated by using a therapy based on modulation of GSH levels in cancer cells.

Salvage surgery after locoregional failure in head and neck carcinoma patients treated with chemoradiotherapy.

With the increasing use of concomitant chemoradiotherapy (CCRT) in the treatment of advanced head and neck carcinoma, surgery has lost ground as the first therapy and is reserved as a salvage treatment in cases of locoregional failure. The objective of our study was to review our experience in patients who had a local or regional recurrence after treatment with CCRT. Thirty-two patients underwent salvage surgery after CCRT: 24 were treated with a local or locoregional resection and 8 patients with a neck dissection only. In patients who had surgery involving the primary location of the tumor, some kind of reconstruction was required in 83% of cases. One or more postoperative complications occurred in nine patients. The median hospital stay was 18.5 days. There was a significant difference in hospital stay in relation to the appearance of surgical complications. Five-year adjusted survival after salvage surgery was 34.2% (CI 95% 13.2-55.2%). Adjusted survival was related to the status of the resection margins and appearance of neck nodes with extracapsular spread in the neck dissection. In conclusion, salvage surgery after CCRT involves extensive resections, requiring reconstruction techniques with regional or microanastomosed free flaps in most cases, achieving acceptable outcomes.

Prenatal vitamin D deficiency does not exacerbate behavioural impairments associated with prenatal ethanol exposure in juvenile male mice.

There is a high prevalence of vitamin D deficiency and exposure to low levels of ethanol in pregnant women. However, there are a paucity of studies that have addressed the impact of both vitamin D deficiency and ethanol exposure on the offspring's vulnerability to neurodevelopmental disorders later in life. The aim of this study was to examine whether the absence of vitamin D during gestation in mice would alter the effects of prenatal exposure to low dose ethanol on the behaviour and dopaminergic gene expression patterns of juvenile mice. Four-week old female C57BL/6J mice were placed on a prenatal vitamin D deficient (PVD) or standard diet for 6 weeks and mated at 10 weeks of age. Females were exposed to either 10%(v/v) ethanol or water between gestational days 0-8 and all were offered water thereafter. We found that blood ethanol concentration in the dams was not affected by maternal diet. Behavioural analyses of the offspring included ultrasonic vocalizations (USV) at postnatal day (P) 7, locomotion and social interaction at P21. The main findings were increased USV calling rate and impaired social interaction in males with prenatal ethanol exposure (PrEE). Gene expression analysis of transcripts involved in dopamine regulation revealed a main effect of ethanol exposure on dopamine- and cyclic adenosine monophosphate- regulated neuronal phosphoprotein (Darpp-32), a main effect of vitamin D diet on Dopamine 2 Receptors (D2R) and a main effect of Sex on Tyrosine Hydroxylase (TH) expression. The combination of PVD-PrEE did not exacerbate the alterations resulting from PVD or PrEE. Despite the limited evidence to support the interaction of PVD and PrEE during the postnatal period, males were more vulnerable than female offspring to the detrimental effects of PrEE. Therefore, based on these studies in mice we suggest that maintenance of optimal vitamin D levels and abstinence from ethanol during pregnancy would reduce risk of later disruption to brain function and behaviour in the offspring.

Thermodynamic and structural characterization of Asn and Ala residues in the disallowed II' region of the Ramachandran plot.

Residue Asn47 at position L1 of a type II' beta-turn of the alpha-spectrin SH3 domain is located in a disallowed region of the Ramachandran plot (phi = 56 +/- 12, psi = -118 +/- 17). Therefore, it is expected that replacement of Asn47 by Gly should result in a considerable stabilization of the protein. Thermodynamic analysis of the N47G and N47A mutants shows that the change in free energy is small (approximately 0.7 kcal/mol; approximately 3 kJ/mol) and comparable to that found when mutating a Gly to Ala in a alpha-helix or beta-sheet. X-ray structural analysis of these mutants shows that the conformation of the beta-turn does not change upon mutation and, therefore, that there is no relaxation of the structure, nor is there any gain or loss of interactions that could explain the small energy change. Our results indicate that the energetic definition of II' region of the Ramachandran plot (phi = 60 +/- 30, psi = -115 +/- 15) should be revised for at least Ala and Asn in structure validation and protein design.

Pharmacokinetic variations of acetaminophen according to liver dysfunction and portal hypertension status.

AIM: To study the pharmacokinetic and metabolism profiles of a single dose of acetaminophen in patients with cirrhosis. METHODS: Oral acetaminophen (1000 mg) was administered to seven healthy subjects and 14 patients with cirrhosis (nine Child-Pugh A or B and five Child-Pugh C grade), being five without and nine with oesophageal varices. Plasma levels of acetaminophen and its metabolites were determined by HPLC. RESULTS: Patients showed a higher mean area under the curve concentration-time (67.4 +/- 22.4 mg h/L vs. 38.8 +/- 4.3 mg h/L; P = 0.01), a lower clearance (166.7 +/- 85.0 mL/min vs. 367.8 +/- 62.5 mL/min; P = 0.01) and higher elimination half-life (3.8 +/- 1.1 h vs. 2.0 +/- 0.4 h; P = 0.01) of acetaminophen than healthy volunteers. The appearance in blood and the urinary excretion of metabolites in patients did not differ from healthy subjects. Absorption profile was faster in patients. Patients with lower mean and systolic arterial pressure had lower AUC of acetaminophen, independently of liver dysfunction stage. CONCLUSIONS: Patients with cirrhosis had a higher AUC and lower clearance of acetaminophen. Acetaminophen attained earlier therapeutic concentrations in patients with oesophageal varices. Mean and systolic arterial pressures were significantly associated with AUC suggesting the importance of the haemodynamic function on the pharmacokinetics of acetaminophen in patients with cirrhosis.

Binding of the anti-inflammatory steroid deflazacort to glucocorticoid receptors in brain and peripheral tissues. In vivo and in vitro studies.

Deflazacort (DFC) is a heterocyclic glucocorticoid with anti-inflammatory activity but with decreased side effects. In this study, we have evaluated the capacity of DFC and other glucocorticoids to reach the central nervous system (CNS) in vivo by measuring changes of [3H]dexamethasone (DEX) binding to glucocorticoid receptors (GR) in vitro. GR occupation was effected by DEX in the cerebral cortex, hippocampus, pituitary, liver and thymus, with DFC showing a similar profile except for the cerebral cortex. In contrast, corticosterone weakly occupied GR in the thymus, pituitary and hippocampus and methyl-prednisolone was active only in peripheral tissues. Furthermore, IC50 for DEX in vitro amounted to 15-17 nM in the hippocampus and liver, whereas IC50 for the active metabolite 21-deacetyl-DFC (21-OH-DFC) was 4 times higher. 21-OH-DFC bound to type II and was absent from type I GR. When tested in equipotent doses based on IC50 analysis, DFC and DEX similarly induced in vivo ornithine decarboxylase activity in hippocampus and liver, although body weight loss after chronic treatment was significantly less for DFC. The results show that DFC distributes on the CNS similarly to DEX, induces ornithine decarboxylase activity but presents less intensive catabolic effects, making it suitable for use as an anti-inflammatory steroid during chronic therapeutic regimes.

Effects of different continium dielectric models in a molecular dynamics and energy minimization study of the antigenic loop of foot-and-mouth disease virus.

This study presents the influence of the dielectric constant on the final structure of the major antigenic loop of the FMDV serotype C. Minimizations have been performed on the nine-residue peptide Ac-TASARGDLA-NHMe, using two expressions for the dielectric constant: a distance-independent (epsilon = kappa), and a distance-dependent (epsilon = kappa *r) forms, and for kappa values from 1 to 10. In addition, kappa = 40 and 80 has also been considered for the constant expression of the dielectrics. The calculations suggest, for both expressions of the dielectrics, that for the sequence under study a value of kappa in the range of 2-6 performs an adequate treatment of the electrostatic interactions. Finally, molecular dynamic simulations at 298 K calculated with an effective dielectric constants of epsilon = 1*r and epsilon = 4*r are compared. The results indicated that a dielectric of epsilon = 4*r provides a peptide fluctuations which are in agreement with recent X-ray diffraction studies.

Conformational study of a nine residue fragment of the antigenic loop of foot-and-mouth disease virus.

The nine-residue peptide Ac-TASARGDLA-NHMe was selected as model peptide in order to understand the conformational features of the antigenic loop of foot-and-mouth disease virus (FMDV). A throughout exploration of the conformational space has been carried out by means of molecular dynamics (MD) and energy minimization. The calculations have been carried out using the AMBER force field. Solvent effects have been included by an effective dielectric constant of epsilon = 4r. The lowest energy conformation presents a secondary structure constituted by an alpha-helix at the N-terminal end followed by two gamma-turns in the central region. The rest of the accessible minima found present also a high tendency to form gamma-turns. Finally, a 100 ps MD trajectory calculation at 298 K suggest a stability of the secondary structure elements of the lowest energy conformation.

[Toxic hepatitis associated with tamoxifen use. A case report and literature review]. / Hepatitis tóxica asociada al uso de tamoxifeno. Presentación de un caso y revisión bibliográfica.

Tamoxifen is an antiestrogenic drug that acts by binding to the estrogen receptor. The drug is used as a co-adjuvant treatment in advanced breast cancer expressing the oestrogen-receptor protein. Clinical trials of tamoxifen have shown its efficacy in reducing mortality and recurrence rates over a five-year treatment. Cases of tamoxifen-associated hepatotoxicity have been described, including cholestasis with or without cytolysis and steatohepatitis. We report the case of a female patient who developed hepatic alterations while undergoing continuous tamoxifen treatment. We also present an overview of similar cases published to date and comment on the advisability of continuing or suppressing this treatment in patients with hepatotoxicity or after a five-year treatment period.

Comportamientos que favorecen la dinámica de reinfestación de Triatomainfestans del Chaco paraguayo / Behaviors that favor reinfestation by Triatoma infestans in the Paraguayan Chaco

El Chaco central paraguayo es unaregión de alto nivel de reinfestación porTriatoma infestans. La población indígena que la habita tiene alta vulnerabilidad por factores culturales y medio ambientales que dificultan acceso y trabajo en la zona. Se propusoconocer factores psicosociales asociadosal proceso de reinfestación para desarrollar tareas de vigilanciacomunitaria. Estudio transversal, con enfoques cualitativo y cuantitativo.Treinta y seis punto siete por ciento (96)de la población de estudio realizómejoras en viviendas; 41,6 % (40)mejoró revoque en paredes. Poblaciónubica al vector en el monte, entre leñas,hojas secas, agujeros de árboles, pozosde topos o tatú; alrededor de animales domésticos, techos de viviendas, gallineros, chiquero de cabras y cerdos.Comprometiendo el traslado pasivo de vinchucas se encontró la recolección deleña 98,5 % (266), del monte, 97,7 %(261) el cambio de lugar de ropas, cajas y comida en las viviendas, 54,7 % (146). Se asoció (p< 0,0005) vivienda mejorada con revoque en paredes yno infestación; viviendas con animales(p< 0,03) e infestación; actitud positivapara eliminar el vector (p<0,04) y no infestación. Comportamientos que comprometen traslado y permanencia de vinchucas fueron acarreo de leña, almacenamiento de comidas yacumulación de ropa y cajas. Paredes revocadas y presencia de animales domésticos se correlacionaron a infestación y actitudes positivas paraeliminación de la vinchuca con viviendas sin reinfestación. Todos ellos sonfactores estratégicos para tareas deprevención y vigilancia con participación comunitaria.

Intrinsic conformational preferences of the Hoechst dye family and their influence of DNA binding.

Quantum mechanical calculations have been used to investigate the molecular conformation of the Hoechst family of DNA-binding dyestuffs. Compounds in which the phenolic substituent adopts either a meta or para position were studied. Two different environments have been considered, which are the gas phase and aqueous solution; the conformation in aqueous solution has been modeled through a self-consistent reaction field strategy. The results clearly indicate that Hoechst dyes do not adopt a planar conformation and that the degree of planarity is controlled by the external environment. A comparison with experimental data reveals that the conformation of Hoechst dyes in the gas phase is similar to that observed in DNA complexes by X-ray crystallography. In aqueous solution, the conformation deviates from planarity more than in the gas phase, since non-bonded interactions with the solvent offset the loss of conjugative interactions. The role of the drug conformation in the binding mechanism with DNA is discussed.

Towards the understanding of the folding of methylene units in the glutamine residue.

The conformational preferences of the methylenic sequence in the side chain of the glutamine residue were investigated by ab initio and semi-empirical quantum mechanical calculations and examination of both the Brookhaven Protein Databank and Cambridge Structural Data Base. The results were analysed on the basis of our previous findings about the folding of methylene groups in aliphatic segments. Both energy calculations and the crystallographic structure of small peptides indicate that methylene units of the glutamine residue tend to fold in a gauche conformation. In contrast, such groups usually adopt an all-trans conformation in proteins due basically to the entropic and solvent contributions. These results have been demonstrated by computing the entropic correction to the free energy and evaluating the solvent effects through SCRF calculations.

The superstructure of chromatin and its condensation mechanism. III: Effect of monovalent and divalent cations X-ray solution scattering and hydrodynamic studies.

Solutions of rat liver and chicken erythrocyte chromatin at different ionic strengths were characterized by synchrotron X-ray solution scattering, ultracentrifugation, density and viscosity measurements. Previous observations on nuclei were extended to rat liver, calf thymus and yeast nuclei. It is shown that with monovalent cations condensation is independent of the nature of the cation whereas with divalent cations there are significant differences related to the preference of base binding over phosphate binding. The consistency of hydrodynamic and scattering results confirm the view that chromatin in solution at low ionic strength has a helix-like superstructure. A survey of X-ray and neutron scattering results in the literature shows that previous interpretations, e.g. in terms of a 10 nm filament, are incompatible with the experimental data at low resolution.

The superstructure of chromatin and its condensation mechanism. II. Theoretical analysis of the X-ray scattering patterns and model calculations.

Model calculations on the superstructure of uncondensed and condensed chromatin are presented. It is found that agreement between the calculated X-ray solution scattering patterns and the experimental observations can be reached with the assumptions that: a) The uncondensed chromatin fibre in solution has a helix-like structure, with a pitch of ca. 33.0 nm, a helical diameter of ca. 20.0 nm and 2.75-3.25 nucleosomes per turn. b) The most condensed state of the chromatin fibre in solution is best represented by a helix-like structure with ca. 2.56 nucleosomes per turn, a pitch of ca. 3.0 nm and a helical diameter of ca. 27.0 nm.

The superstructure of chromatin and its condensation mechanism. I. Synchrotron radiation X-ray scattering results.

Synchroton radiation X-ray scattering experiments have been performed on chicken erythrocyte chromatin fibres over a wide range of ionic conditions and on various states of the fibres (i.e. "native" in solution, in gels and in whole nuclei; chromatin depleted of the H1 (H5) histones and chromatin with bound ethidium bromide). A correlation between the results obtained with the various chromatin preparations provides evidence for a model according to which at low ionic strength the chromatin fibre already possesses a helical superstructure, with a diameter comparable to that of condensed chromatin, held together by the H1(H5) histone. The most significant structural modification undergone upon an increase of the ionic strength is a reduction of the helix pitch, this leads to condensation in a manner similar to the folding of an accordion. The details of this process depend on whether monovalent or divalent cations are used to raise the ionic strength, the latter producing a much higher degree of condensation. Measurements of the relative increase of the mass per unit length indicate that the most condensed state is a helical structure with a pitch around 3.0-4.0 nm. In this paper we give a detailed presentation of the experimental evidence obtained from static and time-resolved scattering experiments, which led to this model.

The superstructure of chromatin and its condensation mechanism. IV. Enzymatic digestion, thermal denaturation, effect of netropsin and distamycin.

Changes in the structure of chicken erythrocyte chromatin fibres at low ionic strength resulting from enzymatic digestion, thermal denaturation and binding of Netropsin and Distamycin were monitored by synchrotron X-ray solution scattering. Digestion with micrococcal nuclease confirms the previous assignment of the 0.05 nm-1 band to an interference between nucleosomes with an average distance of 23 nm. The results of thermal denaturation indicate that above 40 degrees C there is a progressive increase of the internucleosomal distance and that above 60 degrees C the characteristic structure of the chromatin fibre is destroyed. Binding of Netropsin and Distamycin also results in an increase of the internucleosomal distance which can be estimated to correspond to about 0.2 nm/mol.

The three-dimensional structure of Cys-47-modified mouse liver glutathione S-transferase P1-1. Carboxymethylation dramatically decreases the affinity for glutathione and is associated with a loss of electron density in the alphaB-310B region.

The three-dimensional structure of mouse liver glutathione S-transferase P1-1 carboxymethylated at Cys-47 and its complex with S-(p-nitrobenzyl)glutathione have been determined by x-ray diffraction analysis. The structure of the modified enzyme described here is the first structural report for a Pi class glutathione S-transferase with no glutathione, glutathione S-conjugate, or inhibitor bound. It shows that part of the active site area, which includes helix alphaB and helix 310B, is disordered. However, the environment of Tyr-7, an essential residue for the catalytic reaction, remains unchanged. The position of the sulfur atom of glutathione is occupied in the ligand-free enzyme by a water molecule that is at H-bond distance from Tyr-7. We do not find any structural evidence for a tyrosinate form, and therefore our results suggest that Tyr-7 is not acting as a general base abstracting the proton from the thiol group of glutathione. The binding of the inhibitor S-(p-nitrobenzyl)-glutathione to the carboxymethylated enzyme results in a partial restructuring of the disordered area. The modification of Cys-47 sterically hinders structural organization of this region, and although it does not prevent glutathione binding, it significantly reduces the affinity. A detailed kinetic study of the modified enzyme indicates that the carboxymethylation increases the Km for glutathione by 3 orders of magnitude, although the enzyme can function efficiently under saturating conditions.

Solvation in protein folding analysis: combination of theoretical and experimental approaches.

An effort to combine theoretical analyses and protein engineering methods has been made to probe the folding mechanism of SH3 by using Energy Landscape Theory and a phi-value analysis. Particular emphasis was given to core residues and the effect of desolvation during the folding event by replacing the core valines with isosteric threonines. These mutations have the advantage of keeping the core structurally invariant while affecting core stability relative to the unfolded state. Although the valines that form the core appear spatially invariant, the folding kinetics of their threonine mutants varies, indicating their different extent of solvation in the transition-state ensemble. Theoretical studies predicted the distribution of folding kinetics of threonine mutants without previous knowledge of the measured rates. This initial success encourages further investigations of the molecular details behind these macroscopic phenomena and of the role of solvation in the folding mechanism.

Time-resolved synchrotron radiation X-ray solution scattering study of DNA melting.

Time resolved x-ray solution scattering measurements were made during thermal denaturation of DNA from various sources in the temperature range of 20-90 degrees C. Preliminary results on the influence of fragment length, ionic strength, and origin of the DNA on the time course of the scattering are described. Interpretation is based on model calculations of the scattering patterns. The results indicate that, for long DNA fragments at very low ionic strength, the melting process is a continuous phenomenon over the whole temperature range. It is accompanied by a progressive decrease of the radius of gyration of the cross section and of the mass per unit length. For short fragments of 146 base pair nucleosomal core DNA, stiffening of the DNA appears to precede a sharp melting transition.