Leveraging Language Model Multitasking To Predict C-H Borylation Selectivity.

C-H borylation is a high-value transformation in the synthesis of lead candidates for the pharmaceutical industry because a wide array of downstream coupling reactions is available. However, predicting its regioselectivity, especially in drug-like molecules that may contain multiple heterocycles, is not a trivial task. Using a data set of borylation reactions from Reaxys, we explored how a language model originally trained on USPTO_500_MT, a broad-scope set of patent data, can be used to predict the C-H borylation reaction product in different modes: product generation and site reactivity classification. Our fine-tuned T5Chem multitask language model can generate the correct product in 79% of cases. It can also classify the reactive aromatic C-H bonds with 95% accuracy and 88% positive predictive value, exceeding purpose-developed graph-based neural networks.

Neutrophils in Dendritic Cell-Based Cancer Vaccination: The Potential Roles of Neutrophil Extracellular Trap Formation.

Neutrophils have conflicting roles in the context of cancers, where they have been associated with contributing to both anti-tumor and pro-tumor responses. Their functional heterogenicity is plastic and can be manipulated by environmental stimuli, which has fueled an area of research investigating therapeutic strategies targeting neutrophils. Dendritic cell (DC)-based cancer vaccination is an immunotherapy that has exhibited clinical promise but has shown limited clinical efficacy. Enhancing our understanding of the communications occurring during DC cancer vaccination can uncover opportunities for enhancing the DC vaccine platform. There have been observed communications between neutrophils and DCs during natural immune responses. However, their crosstalk has been poorly studied in the context of DC vaccination. Here, we review the dual functionality of neutrophils in the context of cancers, describe the crosstalk between neutrophils and DCs during immune responses, and discuss their implications in DC cancer vaccination. This discussion will focus on how neutrophil extracellular traps can influence immune responses in the tumor microenvironment and what roles they may play in promoting or hindering DC vaccine-induced anti-tumor efficacy.

A very British state capitalism: Variegation, political connections and bailouts during the COVID-19 crisis.

The COVID-19 pandemic has resulted in governments playing increasingly prominent roles as active economic agents. However, state capitalism does not necessarily serve broad developmental purposes, and rather can be directed to supporting sectional and private interests. As the literature on variegated capitalism alerts us, governments and other actors regularly devise fixes in response to a systemic crisis, but the focus, scale, and scope of the interventions vary considerably, according to the constellation of interests. Rapid progress with vaccines notwithstanding, the UK government's response to COVID-19 has been associated with much controversy, not only because of an extraordinarily high death rate, but also because of allegations of cronyism around the granting of government contracts and bailouts. We focus on the latter, investigating more closely who got bailed out. We find that badly affected sectors (e.g. hospitality, transportation) and larger employers were more likely to get bailouts. However, the latter also favored the politically influential and those who had run up debt profligately. Although, as with state capitalism, crony capitalism is most often associated with emerging markets, we conclude that the two have coalesced into a peculiarly British variety, but one that has some common features with other major liberal markets. This might suggest that the eco-systemic dominance of the latter is coming to an end, or, at the least, that this model is drifting towards one that assumes many of the features commonly associated with developing nations.

Kinetic Modeling of API Oxidation: (2) Imipramine Stress Testing.

Gauging the chemical stability of active pharmaceutical ingredients (APIs) is critical at various stages of pharmaceutical development to identify potential risks from drug degradation and ensure the quality and safety of the drug product. Stress testing has been the major experimental method to study API stability, but this analytical approach is time-consuming, resource-intensive, and limited by API availability, especially during the early stages of drug development. Novel computational chemistry methods may assist in screening for API chemical stability prior to synthesis and augment contemporary API stress testing studies, with the potential to significantly accelerate drug development and reduce costs. In this work, we leverage quantum chemical calculations and automated reaction mechanism generation to provide new insights into API degradation studies. In the continuation of part one in this series of studies [Grinberg Dana et al., Mol. Pharm. 2021 18 (8), 3037-3049], we have generated the first ab initio predictive chemical kinetic model of free-radical oxidative degradation for API stress testing. We focused on imipramine oxidation in an azobis(isobutyronitrile) (AIBN)/H2O/CH3OH solution and compared the model's predictions with concurrent experimental observations. We analytically determined iminodibenzyl and desimipramine as imipramine's two major degradation products under industry-standard AIBN stress testing conditions, and our ab initio kinetic model successfully identified both of them in its prediction for the top three degradation products. This work shows the potential and utility of predictive chemical kinetic modeling and quantum chemical computations to elucidate API chemical stability issues. Further, we envision an automated digital workflow that integrates first-principle models with data-driven methods that, when actively and iteratively combined with high-throughput experiments, can substantially accelerate and transform future API chemical stability studies.

Beclin-1 is a novel predictive biomarker for canine cutaneous and subcutaneous mast cell tumors.

Mast cell tumors (MCTs) are the most common skin tumor of the dog, and accurately predicting their clinical behavior is critical in directing patient therapy, as they range from benign lesions to a fatal systemic disease. Grading is useful for prognosis, but it cannot predict the behavior of all MCTs. We hypothesized that biomarker immunolabeling in tumor tissues would correlate with patient morbidity and mortality. A clinically annotated tissue microarray (TMA) of primary, recurrent, and metastatic (to lymph node) canine dermal and subcutaneous MCTs was created. Some dogs whose MCTs were included in the TMA did not receive adjunctive treatment after surgical excision of the MCT, whereas others were treated with one or a combination of chemotherapy, radiation, or oral toceranib. Immunohistochemistry for beclin-1, an autophagy protein, was performed followed by digital image analysis. Beclin-1 immunolabeling was higher in recurrent tumors (mean H-score 110.8) than primary MCTs (mean H-score 73.5), and highest in lymph node metastases (mean H-score 138.5) with a significant difference in means (P < .001). While beclin-1 level was not prognostic, it was strongly predictive for survival after adjunctive treatment; dogs with high beclin-1-expressing tumors showed poorer survival compared to those with low beclin-1-expressing tumors (HR = 5.7, P = .02), especially in Kiupel high-grade tumors (HR = 16.3, P = .01). Beclin-1 immunolabeling was the only significant predictive factor by multivariable analysis (P = .04). These findings may improve our ability to predict the response to adjunctive therapy. Importantly, these data suggest that autophagy inhibitors may be useful in improving response to treatment for dogs with high-grade MCTs.

Proteomic Assessment of Extracellular Vesicles from Canine Tissue Explants as a Pipeline to Identify Molecular Targets in Osteosarcoma: PSMD14/Rpn11 as a Proof of Principle.

Osteosarcoma (OS) is a highly malignant bone tumour that has seen little improvement in treatment modalities in the past 30 years. Understanding what molecules contribute to OS biology could aid in the discovery of novel therapies. Extracellular vesicles (EVs) serve as a mode of cell-to-cell communication and have the potential to uncover novel protein signatures. In our research, we developed a novel pipeline to isolate, characterize, and profile EVs from normal bone and osteosarcoma tissue explants from canine OS patients. Proteomic analysis of vesicle preparations revealed a protein signature related to protein metabolism. One molecule of interest, PSMD14/Rpn11, was explored further given its prognostic potential in human and canine OS, and its targetability with the drug capzimin. In vitro experiments demonstrated that capzimin induces apoptosis and reduces clonogenic survival, proliferation, and migration in two metastatic canine OS cell lines. Capzimin also reduces the viability of metastatic human OS cells cultured under 3D conditions that mimic the growth of OS cells at secondary sites. This unique pipeline can improve our understanding of OS biology and identify new prognostic markers and molecular targets for both canine and human OS patients.

Kinetic Modeling of API Oxidation: (1) The AIBN/H2O/CH3OH Radical "Soup".

Stress testing of active pharmaceutical ingredients (API) is an important tool used to gauge chemical stability and identify potential degradation products. While different flavors of API stress testing systems have been used in experimental investigations for decades, the detailed kinetics of such systems as well as the chemical composition of prominent reactive species, specifically reactive oxygen species, are unknown. As a first step toward understanding and modeling API oxidation in stress testing, we investigated a typical radical "soup" solution an API is subject to during stress testing. Here we applied ab initio electronic structure calculations to automatically generate and refine a detailed chemical kinetics model, taking a fresh look at API oxidation. We generated a detailed kinetic model for a representative azobis(isobutyronitrile) (AIBN)/H2O/CH3OH stress-testing system with a varied cosolvent ratio (50%/50%-99.5%/0.5% vol water/methanol) for 5.0 mM AIBN and representative pH values of 4-10 at 40 °C that was stirred and open to the atmosphere. At acidic conditions, hydroxymethyl alkoxyl is the dominant alkoxyl radical, and at basic conditions, for most studied initial methanol concentrations, cyanoisopropyl alkoxyl becomes the dominant alkoxyl radical, albeit at an overall lower concentration. At acidic conditions, the levels of cyanoisopropyl peroxyl, hydroxymethyl peroxyl, and hydroperoxyl radicals are relatively high and comparable, while, at both neutral and basic pH conditions, superoxide becomes the prominent radical in the system. The present work reveals the prominent species in a common model API stress testing system at various cosolvent and pH conditions, sets the stage for an in-depth quantitative API kinetic study, and demonstrates the usage of novel software tools for automated chemical kinetic model generation and ab initio refinement.

Identifying Conformational Isomers of Organic Molecules in Solution via Unsupervised Clustering.

We present a systematic approach for the identification of statistically relevant conformational macrostates of organic molecules from molecular dynamics trajectories. The approach applies to molecules characterized by an arbitrary number of torsional degrees of freedom and enables the transferability of the macrostates definition across different environments. We formulate a dissimilarity measure between molecular configurations that incorporates information on the characteristic energetic cost associated with transitions along all relevant torsional degrees of freedom. Such metric is employed to perform unsupervised clustering of molecular configurations based on the Fast Search and Find of Density Peaks algorithm. We apply this method to investigate the equilibrium conformational ensemble of Sildenafil, a conformationally complex pharmaceutical compound, in different environments including the crystal bulk, the gas phase, and three different solvents (acetonitrile, 1-butanol, and toluene). We demonstrate that while Sildenafil can adopt more than 100 metastable conformational configurations, only 12 are significantly populated across all of the environments investigated. Despite the complexity of the conformational space, we find that the most abundant conformers in solution are the closest to the conformers found in the most common Sildenafil crystal phase.

Spontaneously occurring melanoma in animals and their relevance to human melanoma.

In contrast to other cancer types, melanoma incidence has been increasing over the last 50 years, and while it still represents less than 5% of all cutaneous malignancies, melanoma accounts for the majority of skin cancer deaths, due to its propensity to metastasise. Whilst melanoma most commonly affects the skin, it can also arise in mucosal surfaces, the eye, and the brain. For new therapies to be developed, a better understanding of the genetic landscape, signalling pathways, and tumour-microenvironmental interactions is needed. This is where animal models are of critical importance. The mouse is the foremost used model of human melanoma. Arguably this is due to its plethora of benefits as a laboratory animal; however, it is important to note that unlike humans, melanocytes are not present at the dermal-epidermal junction in mice and mice do not develop melanoma without genetic manipulation. In contrast, there are numerous reports of animals that spontaneously develop melanoma, ranging from sharks and parrots to hippos and monkeys. In addition, several domesticated and laboratory-bred animals spontaneously develop melanoma or UV-induced melanoma, specifically, fish, opossums, pigs, horses, cats, and dogs. In this review, we look at spontaneously occurring animal 'models' of melanoma and discuss their relevance to the different types of melanoma found in humans. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland..

COVID-19 and digitalization: The great acceleration.

Inspired by burgeoning scholarly interest in the role of digitalization in the COVID-19 pandemic, this paper examines how the COVID-19 pandemic is driving or constraining the digitalization of businesses around the globe. We contend that COVID-19 is "the great accelerator" in fast-tracking the existing global trend towards embracing modern emerging technologies ushering in transformations in lifestyle, work patterns, and business strategies. Thus, COVID-19 has evolved to be a kind of "catalyst" for the adoption and increasing use of digitalization in work organization and the office, alongside presenting foreseen and unforeseen opportunities, challenges, and costs-leading to negative and positive feedback loops. In this article, we develop and advance a conceptual model by linking the different forces for and against digitalization in response to the pandemic. Our analysis indicates that adoption of emerging technologies may be hindered by vested external interests, nostalgia, and employer opportunism, as well as negative effects on employee well-being that undermine productivity, work-life balance, and future of work. Whilst digitalization may bring new opportunities, the process imparts risks that may be hard to mitigate or prepare for. Finally, we draw out the wider theoretical and practical implications of our analysis.

Flow Cytometric Detection of Circulating Osteosarcoma Cells in Dogs.

Osteosarcoma (OSA) is a malignant tumor of middle-aged dogs and adolescent humans. The clinical outcome of OSA has not improved over more than three decades, and dogs typically succumb to metastatic disease within 6 months despite tumor resection through limb amputation and adjuvant chemotherapy. Therefore, undetectable tumor cells with potential to form metastases are present at diagnosis. An assay to identify canine immortalized and primary OSA cells through flow cytometric detection of intracellular collagen 1 (Col I) and osteocalcin was optimized, and applied to blood samples from tumor-bearing dogs for detection of circulating tumor cells (CTCs). Spiking variable number of OSA cells into normal dog blood recovered 50-60% of Col I positive cells with high forward and variable side light scatter. An algorithm to exclude nonviable, doublet, and autofluorescent cells was applied to sequential blood samples from three dogs obtained prior to and after limb amputation, and at approximately, triweekly intervals over 121, 142, and 183 days of chemotherapy, respectively. Dogs had >100 CTC/106 leukocytes prior to amputation, variably frequent CTC during chemotherapy, and an increase up to 4,000 CTC/106 leukocytes within 4 weeks before overt metastases or death. Sorted CTCs were morphologically similar to direct tumor aspirates and positive for Col I. Although preliminary, findings suggest that CTCs are frequent in canine OSA, more numerous than carcinoma CTC in humans, and that an increase in CTC frequency may herald clinical deterioration. This assay may enable enumeration and isolation of OSA CTC for prognostic and functional studies, respectively. © 2019 International Society for Advancement of Cytometry.

Solvation of the Glycyl Radical.

The effect of adding explicit water molecules to the neutral (N) and zwitterionic (Z) forms of the glycyl radical has been examined. The results show that a minimum of three water molecules is required to stabilize the Z radical as a local minimum, with an energy gap of 123 kJ mol-1 between the N and Z forms at this point, in favor of the N form. Increasing the number of water molecules to ∼20 leads to a converged Z-N energy difference of ∼50 kJ mol-1 still in favor of the N form, even though the radical is not considered fully solvated from a structural point of view. Thus, energetic convergence is determined mainly by solvation of the polar functional groups, and a complete coverage of the entire molecule is not necessary. Because aqueous closed-shell glycine exists as a zwitterion while aqueous glycyl radical prefers the neutral form, the conversion between the two necessitates a change along the hydrogen-abstraction reaction pathway. In this regard, the transition structure for α-hydrogen abstraction by the ·OH radical has greater resemblance to glycine than to the glycyl radical. Overall, the barrier for hydrogen abstraction from Z glycine is larger than that from the N isomer, and this might act to provide some protection against radical damage to the free amino acid in the (aqueous) biological environment.

An evaluation of TAZ and YAP crosstalk with TGFß signalling in canine osteosarcoma suggests involvement of hippo signalling in disease progression.

BACKGROUND: Osteosarcoma (OSA) is the most common bone cancer in canines. Both transforming growth factor beta (TGFß) and Hippo pathway mediators have important roles in bone development, stemness, and cancer progression. The role of Hippo signalling effectors TAZ and YAP has never been addressed in canine OSA. Further, the cooperative role of TGFß and Hippo signalling has yet to be explored in osteosarcoma. To address these gaps, this study investigated the prognostic value of TAZ and YAP alone and in combination with pSmad2 (a marker of active TGFß signalling), as well as the involvement of a TGFß-Hippo signalling crosstalk in tumourigenic properties of OSA cells in vitro. An in-house trial tissue microarray (TMA) which contained 16 canine appendicular OSA cases undergoing standard care and accompanying follow-up was used to explore the prognostic role of TAZ, YAP and pSmad2. Published datasets were used to test associations between TAZ and YAP mRNA levels, metastasis, and disease recurrence. Small interfering RNAs specific to TAZ and YAP were utilized in vitro alone or in combination with TGFß treatment to determine their role in OSA viability, proliferation and migration. RESULTS: Patients with low levels of both YAP and pSmad2 when evaluated in combination had a significantly longer time to metastasis (log-rank test, p = 0.0058) and a longer overall survival (log rank test, p = 0.0002). No similar associations were found for TAZ and YAP mRNA levels. In vitro, TAZ knockdown significantly decreased cell viability, proliferation, and migration in metastatic cell lines, while YAP knockdown significantly decreased viability in three cell lines, and migration in two cell lines, derived from either primary tumours or their metastases. The impact of TGFß signaling activation on these effects was cell line-dependent. CONCLUSIONS: YAP and pSmad2 have potential prognostic value in canine appendicular osteosarcoma. Inhibiting YAP and TAZ function could lead to a decrease in viability, proliferation, and migratory capacity of canine OSA cells. Assessment of YAP and pSmad2 in larger patient cohorts in future studies are needed to further elucidate the role of TGFß-Hippo signalling crosstalk in canine OSA progression.

Histologic Grade Does Not Predict Outcome in Dogs with Appendicular Osteosarcoma Receiving the Standard of Care.

Canine appendicular osteosarcoma is an aggressive bone neoplasm that imposes a short survival time. There are several published histologic grading systems for canine osteosarcoma but no universally accepted system. Location within the skeleton and therapy received are both correlated with survival time, but these factors were not always considered when the prognostic value of published grading systems was determined. Our objective was to compare 2 published histologic grading systems in a population of dogs with appendicular osteosarcoma treated with the standard of care for curative intent. Three evaluators graded 85 tumors using 2 histologic grading systems. The relationships between histologic grade as well as individual histologic features and outcome (survival time and disease-free interval) were evaluated using Kaplan-Meier survival functions and a univariate Cox proportional hazards model. Histologic grade, as assigned by any evaluator, did not correlate with outcome. Increased number of mitotic figures per 3 randomly selected 400× microscope fields, as assessed by 1 evaluator, was correlated with both survival time and disease-free interval; this was the only individual histologic feature that was significantly correlated with outcome for any evaluator. These findings cast doubt on the predictive value of routine histologic grading in dogs with appendicular osteosarcoma who receive amputation followed by adjuvant chemotherapy and highlight the need for better tools to predict outcome in canine appendicular osteosarcoma.

AAV vector distribution in the mouse respiratory tract following four different methods of administration.

BACKGROUND: Targeted delivery of gene therapy vectors to the mouse respiratory tract is often performed via intranasal or intratracheal administration; however, there can be a great deal of variability between these methods, which could potentially influence experimental results. Improving the accuracy and precision of lung delivery will not only reduce the number of animals required to detect statistically significant differences, but may reduce the variability of studies from different laboratories. RESULTS: Here we evaluated three different methods of adeno-associated virus (AAV) vector administration to the respiratory tract in mice (intranasal, intubation, and intratracheal injection) and discuss the advantages, challenges, and shortcomings of each. We also present a modified-intranasal delivery technique that is superior to passive administration of vector into the nares of anesthetized supine animals. Transgene expression was consistently visible in the nasal cavity, trachea, and proximal to middle aspect of all lung lobes for all four methods, whereas transgene expression was consistently observed in the most distal aspect of lung lobes only with the intubation and intratracheal injection techniques. AAV vector genome copy numbers in the lung were approximately four-fold lower in mice that received vector via intranasal administration in comparison to the other three methods of vector delivery. The modified intranasal, intubation and intratracheal injection methods of vector administration did not yield statistical differences in AAV vector genome copy numbers in the lung. With regard to reproducibility of vector distribution within and between animals, the modified-intranasal technique was superior. CONCLUSION: Our results show that mode of AAV vector administration to the murine respiratory tract should be selected based on desired target site and skill of the researcher, and that appropriate technique selection may greatly influence experimental outcomes.

Fas receptor expression in germinal-center B cells is essential for T and B lymphocyte homeostasis.

Fas is highly expressed in activated and germinal center (GC) B cells but can potentially be inactivated by misguided somatic hypermutation. We employed conditional Fas-deficient mice to investigate the physiological functions of Fas in various B cell subsets. B cell-specific Fas-deficient mice developed fatal lymphoproliferation due to activation of B cells and T cells. Ablation of Fas specifically in GC B cells reproduced the phenotype, indicating that the lymphoproliferation initiates in the GC environment. B cell-specific Fas-deficient mice also showed an accumulation of IgG1(+) memory B cells expressing high amounts of CD80 and the expansion of CD28-expressing CD4(+) Th cells. Blocking T cell-B cell interaction and GC formation completely prevented the fatal lymphoproliferation. Thus, Fas-mediated selection of GC B cells and the resulting memory B cell compartment is essential for maintaining the homeostasis of both T and B lymphocytes.

Drug-Excipient Interactions in the Solid State: The Role of Different Stress Factors.

Understanding properties and mechanisms that govern drug degradation in the solid state is of high importance to ensure drug stability and safety of solid dosage forms. In this study, we attempt to understand drug-excipient interactions in the solid state using both theoretical and experimental approaches. The model active pharmaceutical ingredients (APIs) under study are carvedilol (CAR) and codeine phosphate (COP), which are known to undergo esterification with citric acid (CA) in the solid state. Starting from the crystal structures of two different polymorphs of each compound, we calculated the exposure and accessibility of reactive hydroxyl groups for a number of relevant crystal surfaces, as well as descriptors that could be associated with surface stabilities using molecular simulations. Accelerated degradation experiments at elevated temperature and controlled humidity were conducted to assess the propensity of different solid forms of the model APIs to undergo chemical reactions with anhydrous CA or CA monohydrate. In addition, for CAR, we studied the solid state degradation at varying humidity levels and also under mechano-activation. Regarding the relative degradation propensities, we found that variations in the exposure and accessibility of molecules on the crystal surface play a minor role compared to the impact of molecular mobility due to different levels of moisture. We further studied drug-excipient interactions under mechano-activation (comilling of API and CA) and found that the reaction proceeded even faster than in physical powder mixtures kept at accelerated storage conditions.

Cross-species models of human melanoma.

Although transformation of melanocytes to melanoma is rare, the rapid growth, systemic spread, as well as the chemoresistance of melanoma present significant challenges for patient care. Here we review animal models of melanoma, including murine, canine, equine, and zebrafish models, and detail the immense contribution these models have made to our knowledge of human melanoma development, and to melanocyte biology. We also highlight the opportunities for cross-species comparative genomic studies of melanoma to identify the key molecular events that drive this complex disease.

Legal Origin and Social Solidarity: The Continued Relevance of Durkheim to Comparative Institutional Analysis.

By using the classic works of Durkheim as a theoretical platform, this research explores the relationship between legal systems and social solidarity. We found that certain types of civil law system, most notably those of Scandinavia, are associated with higher levels of social capital and better welfare state provision. However, we found the relationship between legal system and societal outcomes is considerably more complex than suggested by currently fashionable economistic legal origin approaches, and more in line with the later writings of Durkheim, and, indeed, the literature on comparative capitalisms. Relative communitarianism was strongly affected by relative development, reflecting the complex relationship between institutions, state capabilities and informal social ties and networks.