Preclinical Immunogenicity and Efficacy Studies for Therapeutic Vaccines for Human Papillomavirus-Type-16-Associated Cancer.

The objective of this study was to conduct preclinical immunogenicity and efficacy studies with several therapeutic vaccines for human papillomavirus (HPV)-16-associated cancers expressing the early antigens E5, E6, and E7 with or without E2. The viral oncoproteins were either expressed by themselves as fusion proteins or the fusion proteins were inserted genetically into herpes simplex virus (HSV)-1 glycoprotein D (gD) which, upon binding to the herpes virus entry mediator (HVEM), inhibits an early T cell checkpoint mediated by the B and T cell mediator (BTLA). This, in turn, lowers the threshold for T cell activation and augments and broadens CD8+ T cell responses to the antigens. The fusion antigens were expressed by chimpanzee adenovirus (AdC) vectors. Expression of the HPV antigens within gD was essential for vaccine immunogenicity and efficacy against challenge with TC-1 cells, which express E7 and E6 of HPV-16 but neither E5 nor E2. Unexpectedly, inclusion of E2 increased both CD8+ T cell responses to the other oncoproteins of HPV-16 and the effectiveness of the vaccines to cause the regression of sizable TC-1 tumors.

Cyclocondensation of 2-(α-Diazoacyl)-2H-azirines with Amidines in Diazo Synthesis of Functionalized Naphtho[1,2-d]imidazoles.

A diazo approach toward functionalized naphtho[1,2-d]imidazole derivatives has been developed. It involved a new reaction of arylamidines with 2-(α-diazoacyl)-2H-azirines giving 5-aryl-4-(α-diazoacyl)-1H-imidazoles under mild conditions in good yields. The mechanism of annulation of azirines with amidines is discussed based on DFT calculations. The reaction proceeds in an unusual manner by cleavage of the azirine C-C bond, allowing for the transfer of the aryl substituent from the arylamidine to the proper position of the key intermediate of naphtho[1,2-d]imidazole synthesis. Under thermolysis conditions, 5-aryl-4-(α-diazoacyl)-1H-imidazoles undergo Wolff rearrangement followed by the selective 6π-cyclization of transient ketene to form 3H-naphtho[1,2-d]imidazoles bearing various substituents in the positions 2,3,4,5,7,8,9. Additionally, variation of the substituents at position 5 of naphtho[1,2-d]imidazoles is possible through the formation of triflates and subsequent cross-coupling reactions. One more heterocyclic pharmacophoric skeleton, 3H-furo[3',2':3,4]naphtho[1,2-d]imidazole, was easily constructed from methyl 5-hydroxy-3H-naphtho[1,2-d]imidazole-4-carboxylates in a one-pot mode using O-alkylation with phenacyl bromides followed by base-induced intramolecular acyl substitution at room temperature with high yields.

Acridine-Isoxazole and Acridine-Azirine Hybrids: Synthesis, Photochemical Transformations in the UV/Visible Radiation Boundary Region, and Anticancer Activity.

Easy-to-handle N-hydroxyacridinecarbimidoyl chloride hydrochlorides were synthesized as convenient nitrile oxide precursors in the preparation of 3-(acridin-9/2-yl)isoxazole derivatives via 1,3-dipolar cycloaddition with terminal alkynes, 1,1-dichloroethene, and acrylonitrile. Azirines with an acridin-9/2-yl substituent attached directly or via the 1,2,3-triazole linker to the azirine C2 were also synthesized. The three-membered rings of the acridine-azirine hybrids were found to be resistant to irradiation in the UV/visible boundary region, despite their long-wave absorption at 320-420 nm, indicating that the acridine moiety cannot be used as an antenna to transfer light energy to generate nitrile ylides from azirines for photoclick cycloaddition. The acridine-isoxazole hybrids linked at the C9-C3 or C2-C3 atoms under blue light irradiation underwent the addition of such hydrogen donor solvents, such as, toluene, o-xylene, mesitylene, 4-chlorotoluene, THF, 1,4-dioxane, or methyl tert-butyl ether (MTBE), to the acridine system to give the corresponding 9-substituted acridanes in good yields. The synthesized acridine-azirine, acridine-isoxazole, and acridane-isoxazole hybrids exhibited cytotoxicity toward both all tested cancer cell lines (HCT 116, MCF7, and A704) and normal cells (WI-26 VA4).

Stable 2-Azaallyl Salts as a Bridge between 2H-Azirines and Densely Functionalized 2H-Pyrroles.

An efficient protocol for the synthesis of stable 2-azaallyl anion salts by the reaction of alkyl 2-bromo-2H-azirine-2-carboxylates with trimethylsilyl cyanide/Bu4NF has been developed. The domino reaction proceeds in four steps via the cleavage of the azirine C-C bond to provide the tetrabutylammonium salts of stereochemically pure 2-azaallyl anions having U-configuration relative to the cyano groups. The anions with an ortho-substituted aryl group or styryl group exist as a mixture of two geometrical isomers across the N2-C3 bond. 2-Azaallyl anion salts have been shown to be convenient substrates for the one-pot synthesis of densely functionalized 2H-pyrroles by the alkylation-cyclization sequence.

Divergent Diazo Approach toward Alkyl 5/4-Hydroxy-3H-benzo[e]indole-4/5-carboxylates.

A divergent diazo approach toward alkyl 5/4-hydroxy-3H-benzo[e]indole-4/5-carboxylates has been developed. The reaction of 1,3-diketones with alkyl 2-diazo-3-oxo-3-(2H-azirin-2-yl)propanoates catalyzed by Co(acac)3 or Ni(acac)2 gives various alkyl 3-(1H-pyrrol-2-yl)-2-diazo-3-oxopropanoate in good yields. The latter undergo Wolff rearrangement followed by the 6π-cyclization of transient ketene to form alkyl 5-hydroxy-3H-benzo[e]indole-4-carboxylates bearing various substituents in positions 1, 2, 7, and 8, as well as derivatives of methyl 4-hydroxy-6H-thieno[2,3-e]indole-5-carboxylates and methyl 5-hydroxy-7H-benzo[c]carbazole-6-carboxylate under thermolysis or Rh2(OAc)4 catalysis. Isomeric benzoindoles, alkyl 4-hydroxy-3H-benzo[e]indole-5-carboxylates, have been prepared by Boc-protection of the pyrrole nitrogen of alkyl 3-(1H-pyrrol-2-yl)-2-diazo-3-oxopropanoates followed by an intramolecular formal carbene insertion into the aromatic C-H bond catalyzed by Cu(OTf)2. The hydroxyl group of alkyl 5/4-hydroxy-3H-benzo[e]indole-4/5-carboxylates, through the formation of the corresponding triflates, allows the introduction of various substituents into the 5/4 position of benzo[e]indoles using the cross-coupling reaction and even form a new heterocyclic backbone, benzo[k]pyrrolo[2,3-i]phenanthridine, via a tandem Suzuki reaction/nucleophilic acyl substitution.

Divergent Synthesis of Pyrazolo[1,5-a]pyridines and Imidazo[1,5-a]pyridines via Reagent-Controlled Cleavage of the C-N or C-C Azirine Bond in 2-Pyridylazirines.

The three-membered ring in 2-(2-pyridyl)azirine-2-carboxylic esters and thioesters can undergo selective cleavage of either the N-C2 bond under copper(II) catalysis or the C-C bond under the action of HCl to provide isomeric azirine ring expansion products of pyrazolo[1,5-a]pyridine or imidazo[1,5-a]pyridine series, respectively. Mild catalytic reaction conditions for the formation of pyrazolopyridines make it possible to obtain them directly from 4-bromoisoxazoles by a one-pot, three-stage procedure without isolating the intermediate 2-bromoazirines and 2-(2-pyridyl)azirines.

Triethylamine-Promoted Oxidative Cyclodimerization of 2H-Azirine-2-carboxylates to Pyrimidine-4,6-dicarboxylates: Experimental and DFT Study.

An unprecedented oxidative cyclodimerization reaction of 2H-azirine-2-carboxylates to pyrimidine-4,6-dicarboxylates under heating with triethylamine in air is described. In this reaction, one azirine molecule undergoes formal cleavage across the C-C bond and another across the C=N bond. According to the experimental study and DFT calculations, the key steps of the reaction mechanism include nucleophilic addition of N,N-diethylhydroxylamine to an azirine to form an (aminooxy)aziridine, generation of an azomethine ylide, and its 1,3-dipolar cycloaddition to the second azirine molecule. The crucial condition for the synthesis of pyrimidines is generation of N,N-diethylhydroxylamine in the reaction mixture in a very low concentration, which is ensured by the slow oxidation of triethylamine with air oxygen. Addition of a radical initiator accelerated the reaction and resulted in higher yields of the pyrimidines. Under these conditions, the scope of the pyrimidine formation was elucidated, and a series of pyrimidines was synthesized.

Tandem Cycloaddition of Azides to 3,3-Diaminoacrylonitriles (2-Cyanoacetamidines) and Cornforth-Type Rearrangement as an Approach to 5-Amino-1,2,3-triazole-4-N-substituted Imidamides.

An efficient base-catalyzed, metal-free method for the synthesis of 5-amino-1,2,3-triazole-4-N-sulfonyl- and arylimidamides, directed by the structure of the amidine group, has been developed. It is based on a previously unknown tandem process involving cycloaddition reaction to 3,3-diaminoacrylonitriles (2-cyanoacetamidines) with aryl(alkyl)sulfonyl or aryl azides and Cornforth-type rearrangement. During the reaction optimization, different factors were found to facilitate the title reaction, which include the use of a strong base and N-mono- or N,N'-disubstituted 3,3-diaminoacrylonitriles. The reaction is tolerant to variously N-monosubstituted and N,N'-disubstituted 3,3-diaminoacrylonitriles and to various aryl- and aryl/alkyl sulfonyl azides. The developed method has a broad scope and can be applied to obtain a variety of 5-amino-1,2,3-triazole-4-carbimidamides bearing at the N1 position alkyl, allyl, propargyl, benzyl, cycloalkyl, and heteroaryl substituents and sulfonyl and aryl substituents at the amidine group. Post-cyclization reactions of prepared 5-amino-1,2,3-triazoles with DMF-DMA DMA-DMF leads to 1,2,3-triazolo[4,5-d]pyrimidines, 8-aza purine analogues demonstrating the applicability of the prepared compounds in organic synthesis.

[2 + 2] Cycloaddition/Retro-Electrocyclization/Decarboxylation Reaction Sequence: Access to 4-Aminopyridines from Methylideneisoxazolones and Ynamines.

4-(1-Aminoallylidene)isoxazol-5-ones were synthesized in good yields by the [2 + 2]cycloaddition-retro-electrocyclization reaction of 4-methylideneisoxazol-5(4H)-ones with ynamines. The reaction mechanism and the absence of hetero-Diels-Alder cycloadducts were investigated by DFT calculations. 4-(1-Aminoallylidene)isoxazol-5-ones were found to be convenient substrates for the preparation of 4-aminopyridines under thermal metal-free conditions. The process proceeds via decarboxylative isomerization with cyclic amino groups and accompanied by partial dealkylation of acyclic amino groups. 4-Aminopyridines can also be prepared directly from 4-methylideneisoxazol-5(4H)-ones and ynamines by the one-pot protocol.

Compensatory mechanisms of reduced interhemispheric EEG connectivity during sleep in patients with apnea.

We performed a mathematical analysis of functional connectivity in electroencephalography (EEG) of patients with obstructive sleep apnea (OSA) (N = 10; age: 52.8 ± 13 years; median age: 49 years; male/female ratio: 7/3), compared with a group of apparently healthy participants (N = 15; age: 51.5 ± 29.5 years; median age: 42 years; male/female ratio: 8/7), based on the calculation of wavelet bicoherence from nighttime polysomnograms. Having observed the previously known phenomenon of interhemispheric synchronization deterioration, we demonstrated a compensatory increase in intrahemispheric connectivity, as well as a slight increase in the connectivity of the central and occipital areas for high-frequency EEG activity. Significant changes in functional connectivity were extremely stable in groups of apparently healthy participants and OSA patients, maintaining the overall pattern when comparing different recording nights and various sleep stages. The maximum variability of the connectivity was observed at fast oscillatory processes during REM sleep. The possibility of observing some changes in functional connectivity of brain activity in OSA patients in a state of passive wakefulness opens up prospects for further research. Developing the methods of hypnogram evaluation that are independent of functional connectivity may be useful for implementing a medical decision support system.

A new view on the mechanism of UV photodegradation of the tricyclic antidepressant carbamazepine in aqueous solutions.

Mechanism of direct UV photolysis of the tricyclic antidepressant carbamazepine (CBZ) at neutral pH was revealed by a combination of nanosecond laser flash photolysis, steady-state photolysis combined with high resolution LC-MS and DFT quantum-chemical calculations. The detection of short-lived intermediates and the detailed identification of final products were performed for the first time. The quantum yield of CBZ photodegradation (282 nm) is about 0.1% and 0.18% in air-equilibrated and argon-saturated solutions. The primary stage is photoionization with the formation of CBZ cation radical followed by a rapid nucleophilic attack by a solvent molecule. The primary photoproducts are 10-oxo-9-hydro-carbamazepine, 9-formylacridine-10(9H)-carboxamide (a result of ring contraction) and various isomers of hydroxylated CBZ. Prolonged irradiation results to accumulation of acridine derivatives, which should lead to an increase of the toxicity of photolyzed CBZ solutions. The obtained results may be important for understanding the fate of tricyclic antidepressants in processes of UVC disinfection and in natural waters under action of sunlight.

Gold vs Light: Chemodivergent Reactivity of Diazoesters toward 2H-Azirine-2-carboxylic Acids.

An orthogonal reactivity of diazo compounds toward azirine-2-carboxylic acids, switching with the reaction conditions, is demonstrated. A gold-catalyzed reaction is N-selective and produces 1,3-oxazin-6-ones, whereas a blue light activation leads to O-H insertion products, azirine-2-carboxylic esters. The observed chemodivergence is explained by the metal-bound and metal-free carbenes exhibiting different electronic properties in these reactions. In addition, a high antibacterial potential of the 1,3-oxazin-6-ones synthesized is shown.

A Complex of Marine Geophysical Methods for Studying Gas Emission Process on the Arctic Shelf.

The Russian sector of the arctic shelf is the longest in the world. Quite a lot of places of massive discharge of bubble methane from the seabed into the water column and further into the atmosphere were found there. This natural phenomenon requires an extensive complex of geological, biological, geophysical, and chemical studies. This article is devoted to aspects of the use of a complex of marine geophysical equipment applied in the Russian sector of the arctic shelf for the detection and study of areas of the water and sedimentary strata with increased saturation with natural gases, as well as a description of some of the results obtained. This complex contains a single-beam scientific high-frequency echo sounder and multibeam system, a sub-bottom profiler, ocean-bottom seismographs, and equipment for continuous seismoacoustic profiling and electrical exploration. The experience of using the above equipment and the examples of the results obtained in the Laptev Sea have shown that these marine geophysical methods are effective and of particular importance for solving most problems related to the detection, mapping, quantification, and monitoring of underwater gas release from the bottom sediments of the shelf zone of the arctic seas, as well as the study of upper and deeper geological roots of gas emission and their relationship with tectonic processes. Geophysical surveys have a significant performance advantage compared to any contact methods. The large-scale application of a wide range of marine geophysical methods is essential for a comprehensive study of the geohazards of vast shelf zones, which have significant potential for economic use.

Metal carbonyl mediated rearrangement of 5-(2-oxoalkyl)-1,2,4-oxadiazoles: synthesis of fully substituted pyrimidines.

Variously substituted ethyl 6-oxo-1,6-dihydropyrimidine-5-carboxylates can be easily prepared by a metal carbonyl mediated rearrangement of ethyl 3-oxo-2-(1,2,4-oxadiazol-5-yl)propanoates. The irradiation of a mixture of oxadiazoles and Fe(CO)5 in wet solvents with a 365 nm LED at room temperature for 2 h followed by heating at 80 °C for 2 h gives pyrimidines in up to 90% yield. This procedure enables the preparation of 6-oxo-1,6-dihydropyrimidine-5-carboxylates with various aryl substituents at the C2 and alkyl or aryl substituents at the C4 position. 1-(1,2,4-Oxadiazol-5-yl)propan-2-ones analogously give 6-methylpyrimidin-4(3H)-ones, albeit in lower yields. Ethyl 6-oxo-1,6-dihydropyrimidine-5-carboxylates can be easily modified at the C6 position by bromination followed by cross-coupling reactions to give pyrimidine-5-carboxylates with pyridyl, amino and ethynyl substituents.

The Objective Assessment of Event-Related Potentials: An Influence of Chronic Pain on ERP Parameters.

The article presents an original method for the automatic assessment of the quality of event-related potentials (ERPs), based on the calculation of the coefficient ε, which describes the compliance of recorded ERPs with some statistically significant parameters. This method was used to analyze the neuropsychological EEG monitoring of patients suffering from migraines. The frequency of migraine attacks was correlated with the spatial distribution of the coefficients ε, calculated for EEG channels. More than 15 migraine attacks per month was accompanied by an increase in calculated values in the occipital region. Patients with infrequent migraines exhibited maximum quality in the frontal areas. The automatic analysis of spatial maps of the coefficient ε demonstrated a statistically significant difference between the two analyzed groups with different means of migraine attack numbers per month.

Heterologous chimpanzee adenovirus vector immunizations for SARS-CoV-2 spike and nucleocapsid protect hamsters against COVID-19.

Available COVID-19 vaccine only provide protection for a limited time due in part to the rapid emergence of viral variants with spike protein mutations, necessitating the generation of new vaccines to combat SARS-CoV-2. Two serologically distinct replication-defective chimpanzee-origin adenovirus (Ad) vectors (AdC) called AdC6 and AdC7 expressing early SARS-CoV-2 isolate spike (S) or nucleocapsid (N) proteins, the latter expressed as a fusion protein within herpes simplex virus glycoprotein D (gD), were tested individually or as a mixture in a hamster COVID-19 SARS-CoV-2 challenge model. The S protein expressing AdC (AdC-S) vectors induced antibodies including those with neutralizing activity that in part cross-reacted with viral variants. Hamsters vaccinated with the AdC-S vectors were protected against serious disease and showed accelerated recovery upon SARS-CoV-2 challenge. Protection was enhanced if AdC-S vectors were given together with the AdC vaccines that expressed the gD N fusion protein (AdC-gDN). In contrast hamsters that just received the AdC-gDN vaccines showed only marginal lessening of symptoms compared to control animals. These results indicate that immune response to the N protein that is less variable than the S protein may potentiate and prolong protection achieved by the currently used S protein based genetic COVID-19 vaccines.

Examining time-frequency mechanisms of full-fledged deep sleep development in newborns of different gestational age in the first days of their postnatal development.

Early age-related changes in EEG time-frequency characteristics during the restful sleep of newborns of different gestational ages result in the development of conventional EEG signs of deep sleep already during the first postnatal week of their life. Allocating newborns to different groups based on their gestational age and duration of postnatal period allowed demonstrating substantial intergroup differences in brain activity during sleep and wakefulness, along with significant variability in the time-frequency characteristics of brain activity. The process of conventional deep sleep development in infants born prior to the week 35 of gestation is associated with an increase in the power of alpha activity in the sensorimotor cortex of the brain.

Effects of Daily Peat Smoke Exposure on Present and Next Generations.

This study aimed to follow the neurotoxic effect of peat smoke on adult outbred rats and its influence on central nervous system (CNS) parameters in first-generation offspring. Under experimental conditions, exposure to peat smoke was carried out on adult male Wistar rats for 24 h. After the end of the exposure, an open field test (OFT), electroencephalography (EEG), and histological analysis of the testes and brains of smoke-exposed males were performed, after which they were mated with intact females to obtain F1 offspring. Stillbirth, neonatal mortality, and body weight at 4, 7, 14, and 21 postnatal days, as well as behavior in the OFT and EEG parameters during puberty (3 months), were assessed. The results of the examination of F0 males showed a significant increase in motor activity and anxiety in the open field test and a violation of EEG parameters. Histopathologically, peat smoke caused a sharp increase in shadow cells (homogeneous cells with pale-stained cytoplasm, in which the cell and nuclear membranes are not visualized) and degeneratively altered neurons in the brain; we found no changes in the testicles. Peat smoke exposure during preconception did not affect neonatal mortality and weight gain in F1 offspring. Adult females born to peat-smoke-exposed males showed an increase in locomotor activity, and the behavior of adult F1 males did not differ from the control. In F1 males, a statistically significant increase in slow-wave activity indices in the delta band was observed.

Diazo Strategy for Intramolecular Azirine Ring Expansion: Rh(II)-Catalyzed Synthesis of 2-Hydroxy-3-oxo-2,3-dihydro-1H-pyrrole-2-carboxylates.

A diazo strategy for the intramolecular azirine ring expansion was developed. Azirinyl-substituted diazodicarbonyl compounds, prepared from diazoacetylazirines, were converted in excellent yields to 2-hydroxy-3-oxo-2,3-dihydro-1H-pyrrole-2-carboxylates under Rh catalysis in the presence of water. According to DFT calculations, the formation of only pyrrolinone derivatives and the absence of O-H insertion products of Rh carbene into water are due to the low Gibbs free energy of the transition state for the concerted opening of the azirine ring and recyclization to pyrrolone in intermediate Rh complexes.

Copper(II)-Catalyzed (3+2) Cycloaddition of 2H-Azirines to Six-Membered Cyclic Enols as a Route to Pyrrolo[3,2-c]quinolone, Chromeno[3,4-b]pyrrole, and Naphtho[1,8-ef]indole Scaffolds.

A method for the [2+3] pyrroline annulation to the six-membered non-aromatic enols using 3-aryl-2H-azirines as annulation agents is developed in the current study. The reaction proceeds as a formal (3+2) cycloaddition via the N1-C2 azirine bond cleavage and is catalyzed by both Cu(II) and Cu(I) compounds. The new annulation method can be applied to prepare pyrrolo[3,2-c]quinoline, chromeno[3,4-b]pyrrole, and naphtho[1,8-ef]indole derivatives in good to excellent yields from enols of the quinolin-2-one, 2H-chromen-2-one, and 1H-phenalen-1-one series.