[3+2] click chemistry approach to tetrazine containing polymers

We report a [3+2] cycloaddition using 3,6-bis-propargyloxy-1,2,4,5-tetrazine and azides to synthesize energetic polymers containing 1,2,4,5-tetrazine within the scaffold. This work also includes [3+2] cycloaddition to crosslink azide containing glycidyl azide polymer (GAP). These reactions provide pathways for incorporation of 1,2,4,5-tetrazine into novel energetic materials using click-chemistry and provide an alternative polymer curing approach.

We report a [3+2] cycloaddition using 3,6-bis-propargyloxy-1,2,4,5-tetrazine and azides to synthesize energetic polymers containing 1,2,4,5-tetrazine within the scaffold.

Due to their versatility and unique chemical and physical properties, tetrazines have found utility in organic solar cells,^1,2 sensors,^3,4 on/off fluorescence^5–8 and energetic materials applications.^9,10 A significant increase in 1,2,4,5-tetrazine chemistry synthesis has also occurred over the past several years.^11–14 A number of investigations have focused in large part on bioorthogonal, inverse electron demand hetero-Diels–Alder reactions with 1,2,4,5-tetrazines for numerous applications, such as in situ synthesis of fluorogenic probes for live cell imaging, and near infrared fluorogenic probes.⁸ Unfortunately, the inverse electron demand hetero-Diels–Alder reaction of 1,2,4,5-tetrazines is not a promising pathway in energetic materials chemistry as a concomitant reduction in the inherent energy of the molecule results during these types of transformations due to the loss of N₂. Thus alternative pathways for the incorporation of 1,2,4,5-tetrazine into novel energetic materials are warranted.One very interesting possibility for the preparation of new energetics is to employ a [3+2] cycloaddition (click-chemistry) strategy. In this case, a Huisgen 1,3-dipolar cycloaddition between 1,3-dipoles such as azides, and 1,2-dipolarophiles such as acetylenes could be incorporated onto the tetrazine ring system at the 3 and 6 positions either directly or through linkers of varying length. The challenge presented with this approach is the possibility for the 1,2,4,5-tetrazine system to undergo competitive inverse electron demand [4+2] hetero-Diels–Alder chemistry with acetylene functional groups. Indeed, Rusinov and co-workers found that the reaction of but-3-yn-1-ol with 3,6-bis(3,5-dimethylpyrazol-1-yl)-1,2,4,5-tetrazine (1) in the presence of triethylamine in acetonitrile led to the two products 2 and 3 (5 : 1) as shown in Scheme 1.¹⁵ These two products both are a result of inverse electron demand hetero-Diels–Alder processes. Changing the leaving group from 3,5-dimethylpyrazole to imidazole led to the exclusive isolation of the bicyclic product 2.Open in a separate windowScheme 1But-3-yn-1-ol reaction with 1 in the presence of TEA^.While these literature results were not encouraging, we decided to pursue the synthesis of 3,6-bispropargyloxy-1,2,4,5-tetrazine (4). We found that shortening the distance between the alcohol group and the acetylene moiety allowed for nucleophilic addition to proceed exclusively with no evidence of inter- or intramolecular hetero-Diels–Alder cyclization occurring. Thus propargyl alcohol reacted with 1 in the presence of potassium carbonate to provide 4 in excellent yield (Scheme 2). Ultimately, we were able to perform the reaction neat, using propargyl alcohol as the solvent, although only in minimal amounts.Open in a separate windowScheme 2The synthesis of 4.To confirm the structure of 4, X-ray quality crystals were obtained from slow diffusion of diethyl ether into a solution of 4 in acetone. The compound was found to have the monoclinic crystal system with a P2₁/c space group.⁷ It displayed a density of 1.46 g cm⁻³ at 293 K (see ESI^†). The Ortep representation of the crystal structure is displayed in Fig. 1. Overall, the transformation resulting in 4 is unprecedented and allowed for a “click” chemical approach to make energetic, tetrazine containing polymers.Open in a separate windowFig. 1Molecular conformation of 4, nonhydrogen atoms are shown as 50% probability displacement ellipsoids. Ellipsoids of 4 shown at 50% probability.With 4 in hand, we attempted to perform the Huisgen cycloaddition reaction with azide containing materials employing the Cu(i)-catalyzed azide/alkyne cycloaddition approach (CuAAC) to accomplish prepolymer curing, polymerization and self-polymerization.⁸ We first investigated a [3+2] cross-linking strategy using 4 as a curing agent for azide containing prepolymers, such as the glycidyl azide (GAP) prepolymer. A similar approach has been described with other polyacetylene compounds.⁹ Typically GAP is cured using a urethane crosslinking approach, but this method presents challenges, such as sensitivity to moisture and the use of reactive isocyanates. A [3+2] curing approach with a high energy cross linker could provide higher energy GAP polymers. Initial results found that 5 wt% 4 was successful at curing the GAP prepolymer (5) in the presence of a catalytic amount of CuCl, to produce polymer 6 (Scheme 3).Open in a separate windowScheme 3Curing of glycidyl azide prepolymer.These results were expanded on to include a curing study of GAP (5) and GAP plasticizer (GAPp, 5p) with varying weight percentages of 4 in the presence of a copper halide species to produce polymer 6 and 6p, respectively. The GAP (also called GAP 5527-Polyol) is a high molecular weight prepolymer with a hydroxyl equivalent between 2.5–3 used in urethane cures, while the GAPp is a lower viscosity prepolymer with an average MW of 700, and a hydroxyl equivalent close to 1. As the click-chemistry cure for these GAP pre-polymers is independent of hydroxyl terminal group content, the two materials were utilized to evaluate rheological properties over the course of reaction. Rheometry measurements were conducted on samples containing 3% of 4 for both polymers (see ESI^†). The transition from liquid to solid, or gel point, for 6 occurred at roughly 83 minutes compared to 6p at about 4 hours.After demonstrating that 4 could be used to cure GAP, we turned our attention to polymerization studies of 4 with diazide compounds. One such material was 2,2-bisazidomethyl-1,3-propanediol (7).¹⁶ Since 7 is a liquid, we performed the cycloaddition neat by thoroughly mixing 4, 7 with a catalytic amount of a copper halide (CuCl or CuI). Heating for several hours at 60 °C led to the polymerized product (8) (Scheme 4). Similarly, when 4 was reacted with 2-hydroxy-1,3-diazidopropane (9)^17,18 in the presence of a catalytic amount of CuCl, the desired polymerization product (10) was observed (Scheme 4).Open in a separate windowScheme 4Polymerization of 4 with diazides.Polymers 8 and 10 were found to have molecular weights of 3602 ± 136.71 g mol⁻¹ and 6796 ± 132.17 g mol⁻¹, respectively. As expected, Fourier transform infrared spectroscopy (FT-IR) comparisons of 4 to both polymers showed significant reduction in the intensity of the terminal acetylene stretch at 3261 cm⁻¹ (C–H) and 2126 cm⁻¹ (C C) (see ESI^†).Further studies for single compound self-polymerization were explored by incorporation of the azide group and an acetylene group on the same tetrazine ring. We began with the reaction of propargyl alcohol with 3,6-dichloro-1,2,4,5-tetrazine (11) in the presence of 2,4,6-collidine, using tert-butyl methyl ether as a solvent. This provided 12 in good yield (Scheme 5). Subsequent treatment of 12 with sodium azide in acetone provided the 3-azido-6-propargyloxy-1,2,4,5-tetrazine (13).¹⁰ This material melts at 50 °C and begins to polymerize if left in the melt for several hours to provide polymer 14. Unfortunately, the extreme sensitivity of 13 precluded further studies on the properties of the resultant polymer.Open in a separate windowScheme 5Synthesis of 13 and subsequent thermal polymerization to 14.Calculations for compounds 4, 7, and 9 are provided in 19–21Theoretical calculations

The influence of ambient cure chemistry and stoichiometry on epoxy coating surfaces

The surface properties of epoxy resin coatings influence their function as substrates for subsequent coats. Variation in ambient cure conditions (temperature and relative humidity, RH), stoichiometry (ratio of epoxy: amine) and delay time between epoxy component mixing and film casting (“induction time”) significantly altered the surface properties of ambient cured epoxy resin coatings (Dow Epoxy Novolac D.E.N. 431, resorcinol diglycidyl ether and 4,4-diaminodicyclohexylmethane). Gravimetric analysis showed that increasing induction time significantly reduced surface layer formation (carbamation) of cured epoxy resin coatings at 80% RH but had no measurable effect at 40% RH and below. RMS surface roughness increased with increasing RH and decreased with increasing induction time and ambient cure temperature, at two stoichiometric extremes. However, the net change in surface area arising from these conditions was not sufficient to significantly alter the equilibrium contact angles or wetting regime. We conclude that the observed significant variation in surface wettability was more likely to depend on variation in surface chemistry than roughness; stoichiometry was the variable which most significantly influenced surface wettability, average void volume and fractional free volume, while cure temperature significantly influenced the extent of cure at both stoichiometries. Off-stoichiometry formulation and elevated ambient cure temperature significantly increased system average void volume while fractional free volume decreased, which may be significant for the barrier properties of the final coating.

The hydrophilicity of epoxy resin coatings is most sensitive to humidity and induction time. Stoichiometry (amine content) has some effect on surface and strongly influences bulk T_g/void volume along with cure temperature.     

HDACi promotes inflammatory remodeling of the tumor microenvironment to enhance epitope spreading and antitumor immunity

Adoptive cell therapy (ACT) with tumor-specific memory T cells has shown increasing efficacy in regressing solid tumors. However, tumor antigen heterogeneity represents a longitudinal challenge for durable clinical responses due to the therapeutic selective pressure for immune escape variants. Here, we demonstrated that delivery of the class I histone deacetylase inhibitor MS-275 promoted sustained tumor regression by synergizing with ACT in a coordinated manner to enhance cellular apoptosis. We found that MS-275 altered the tumor inflammatory landscape to support antitumor immunoactivation through the recruitment and maturation of cross-presenting CD103⁺ and CD8⁺ DCs and depletion of Tregs. Activated endogenous CD8⁺ T cell responses against nontarget tumor antigens were critically required for the prevention of tumor recurrence. Importantly, MS-275 altered the immunodominance hierarchy by directing epitope spreading toward the endogenous retroviral tumor–associated antigen p15E. Our data suggest that MS-275 in combination with ACT multimechanistically enhanced epitope spreading and promoted long-term clearance of solid tumors.     

An Anterior Cruciate Ligament (ACL) Injury Risk Screening and Reduction Program for High School Female Athletes: A Pilot Study

BackgroundAnterior cruciate ligament (ACL) injury causes physical, mental, and financial burdens. Therefore, it is imperative to screen, identify, and educate athletes who are at high-risk. The combination of screening and education could identify those at risk and potentially reduce future injuries.PurposeThe purpose was to conduct a feasible community pre-season screening program for high school female athletes for the presence of known modifiable risk factors that predispose them to sustaining a non-contact ACL injury.Study DesignNon-experimental prospective studyMethodsA convenience sample of 15 healthy female athletes were recruited from local high schools, consisting of 11 soccer players and four basketball players.  A pre-season screening program was designed encompassing four stations that addressed modifiable neuromuscular and biomechanical risk factors including range of motion (ROM), jump-landing technique, strength, and balance. Athletes were categorized into high-risk versus low-risk groups based on cutoff scores previously established in the literature.ResultsEvery athlete met the high-risk cutoff score for at least one extremity during the ROM screening, and some met high-risk cutoff scores for more than one ROM. Out of all four categories tested, lower extremity ROM demonstrated the greatest deficits.ConclusionThis study identified athletes as having multiple modifiable risk factors that can be addressed with training and exercises. This supports implementing a pre-season program aimed at screening for injury risk factors.Level of EvidenceLevel 3     

Correction to “Retrospective analysis of anti-inflammatory therapies during the first wave of COVID-19 at a community hospital”

Correction to: “Iglesias JI et al. Retrospective analysis of anti-inflammatory therapies during the first wave of COVID-19 at a community hospital. World J Crit Care Med 2021 Sep 9; 10(5): 244-259. DOI: 10.5492/wjccm.v10.i5.244. PMID: 34616660; PMCID: PMC8462025.” In this article, corrections were made to Tables.     

The toxicity of gossypol to the male rat

When (±) gossypol acetic acid was administered to male Sprague-Dawley rats for 26 weeks, the most significant toxicological finding was marked suppression of body weight gain in rats receiving 25 mg/kg per day. Minor biochemical changes were noted at this dosage level. Terminal studies showed 6 out of 20 rats receiving 25 mg/kg per day to have varying degrees of testicular pathology. Five mg/kg per day was shown to be a “no effect” level.     

REACT: Rapid Evaluation Assessment of Clinical Reasoning Tool

IntroductionClinical reasoning encompasses the process of data collection, synthesis, and interpretation to generate a working diagnosis and make management decisions. Situated cognition theory suggests that knowledge is relative to contextual factors, and clinical reasoning in urgent situations is framed by pressure of consequential, time-sensitive decision-making for diagnosis and management. These unique aspects of urgent clinical care may limit the effectiveness of traditional tools to assess, teach, and remediate clinical reasoning.MethodsUsing two validated frameworks, a multidisciplinary group of clinicians trained to remediate clinical reasoning and with experience in urgent clinical care encounters designed the novel Rapid Evaluation Assessment of Clinical Reasoning Tool (REACT). REACT is a behaviorally anchored assessment tool scoring five domains used to provide formative feedback to learners evaluating patients during urgent clinical situations. A pilot study was performed to assess fourth-year medical students during simulated urgent clinical scenarios. Learners were scored using REACT by a separate, multidisciplinary group of clinician educators with no additional training in the clinical reasoning process. REACT scores were analyzed for internal consistency across raters and observations.ResultsOverall internal consistency for the 41 patient simulations as measured by Cronbach’s alpha was 0.86. A weighted kappa statistic was used to assess the overall score inter-rater reliability. Moderate reliability was observed at 0.56.DiscussionTo our knowledge, REACT is the first tool designed specifically for formative assessment of a learner’s clinical reasoning performance during simulated urgent clinical situations. With evidence of reliability and content validity, this tool guides feedback to learners during high-risk urgent clinical scenarios, with the goal of reducing diagnostic and management errors to limit patient harm.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11606-022-07513-5.     

Primary adrenal insufficiency

Primary adrenal insufficiency is a potentially life‐threatening condition. We report a case of a 49‐year‐old female patient who presented to the hospital for evaluation of dizziness, nausea, and vomiting. Darkening of the palmar creases and tongue was noted. The adrenocorticotropic hormone stimulation test confirmed the diagnosis of adrenal insufficiency.

The combination of darkening of the palmar creases, dark tongue, and hyponatremia merits high suspicion and appropriate diagnostic workup for primary adrenal insufficiency due to Addison''s disease.