Development of Non-Natural Flavanones as Antimicrobial Agents

With growing concerns over multidrug resistance microorganisms, particularly strains of bacteria and fungi, evolving to become resistant to the antimicrobial agents used against them, the identification of new molecular targets becomes paramount for novel treatment options. Recently, the use of new treatments containing multiple active ingredients has been shown to increase the effectiveness of existing molecules for some infections, often with these added compounds enabling the transport of a toxic molecule into the infecting species. Flavonoids are among the most abundant plant secondary metabolites and have been shown to have natural abilities as microbial deterrents and anti-infection agents in plants. Combining these ideas we first sought to investigate the potency of natural flavonoids in the presence of efflux pump inhibitors to limit Escherichia coli growth. Then we used the natural flavonoid scaffold to synthesize non-natural flavanone molecules and further evaluate their antimicrobial efficacy on Escherichia coli, Bacillus subtilis and the fungal pathogens Cryptococcus neoformans and Aspergillus fumigatus. Of those screened, we identified the synthetic molecule 4-chloro-flavanone as the most potent antimicrobial compound with a MIC value of 70 µg/mL in E. coli when combined with the inhibitor Phe-Arg-ß-naphthylamide, and MICs of 30 µg/mL in S. cerevesiae and 30 µg/mL in C. neoformans when used alone. Through this study we have demonstrated that combinatorial synthesis of non-natural flavonones can identify novel antimicrobial agents with activity against bacteria and fungi but with minimal toxicity to human cells

Immunization with Radiation-Attenuated Plasmodium berghei Sporozoites Induces Liver cCD8α+DC that Activate CD8+T Cells against Liver-Stage Malaria

Immunization with radiation (γ)-attenuated Plasmodia sporozoites (γ-spz) confers sterile and long-lasting immunity against malaria liver-stage infection. In the P. berghei γ-spz model, protection is linked to liver CD8+ T cells that express an effector/memory (TEM) phenotype, (CD44hiCD45RBloCD62Llo ), and produce IFN-γ. However, neither the antigen presenting cells (APC) that activate these CD8+ TEM cells nor the site of their induction have been fully investigated. Because conventional (c)CD8α+ DC (a subset of CD11c+ DC) are considered the major inducers of CD8+ T cells, in this study we focused primarily on cCD8α+ DC from livers of mice immunized with Pb γ-spz and asked whether the cCD8α+ DC might be involved in the activation of CD8+ TEM cells. We demonstrate that multiple exposures of mice to Pb γ-spz lead to a progressive and nearly concurrent accumulation in the liver but not the spleen of both the CD11c+NK1.1− DC and CD8+ TEM cells. Upon adoptive transfer, liver CD11c+NK1.1− DC from Pb γ-spz-immunized mice induced protective immunity against sporozoite challenge. Moreover, in an in vitro system, liver cCD8α+ DC induced naïve CD8+ T cells to express the CD8+ TEM phenotype and to secrete IFN-γ. The in vitro induction of functional CD8+ TEM cells by cCD8α+ DC was inhibited by anti-MHC class I and anti-IL-12 mAbs. These data suggest that liver cCD8α+ DC present liver-stage antigens to activate CD8+ TEM cells, the pre-eminent effectors against pre-erythrocytic malaria. These results provide important implications towards a design of anti-malaria vaccines

Why Functional Pre-Erythrocytic and Bloodstage Malaria Vaccines Fail: A Meta-Analysis of Fully Protective Immunizations and Novel Immunological Model

Background: Clinically protective malaria vaccines consistently fail to protect adults and children in endemic settings, and at best only partially protect infants. Methodology/Principal Findings: We identify and evaluate 1916 immunization studies between 1965-February 2010, and exclude partially or nonprotective results to find 177 completely protective immunization experiments. Detailed reexamination reveals an unexpectedly mundane basis for selective vaccine failure: live malaria parasites in the skin inhibit vaccine function. We next show published molecular and cellular data support a testable, novel model where parasite-host interactions in the skin induce malaria-specific regulatory T cells, and subvert early antigen-specific immunity to parasite-specific immunotolerance. This ensures infection and tolerance to reinfection. Exposure to Plasmodium-infected mosquito bites therefore systematically triggers immunosuppression of endemic vaccine-elicited responses. The extensive vaccine trial data solidly substantiate this model experimentally. Conclusions/Significance: We conclude skinstage-initiated immunosuppression, unassociated with bloodstage parasites, systematically blocks vaccine function in the field. Our model exposes novel molecular and procedural strategies to significantly and quickly increase protective efficacy in both pipeline and currently ineffective malaria vaccines, and forces fundamental reassessment of central precepts determining vaccine development. This has major implications fo

The impact of surgical delay on resectability of colorectal cancer: An international prospective cohort study

AIM: The SARS-CoV-2 pandemic has provided a unique opportunity to explore the impact of surgical delays on cancer resectability. This study aimed to compare resectability for colorectal cancer patients undergoing delayed versus non-delayed surgery. METHODS: This was an international prospective cohort study of consecutive colorectal cancer patients with a decision for curative surgery (January-April 2020). Surgical delay was defined as an operation taking place more than 4 weeks after treatment decision, in a patient who did not receive neoadjuvant therapy. A subgroup analysis explored the effects of delay in elective patients only. The impact of longer delays was explored in a sensitivity analysis. The primary outcome was complete resection, defined as curative resection with an R0 margin. RESULTS: Overall, 5453 patients from 304 hospitals in 47 countries were included, of whom 6.6% (358/5453) did not receive their planned operation. Of the 4304 operated patients without neoadjuvant therapy, 40.5% (1744/4304) were delayed beyond 4 weeks. Delayed patients were more likely to be older, men, more comorbid, have higher body mass index and have rectal cancer and early stage disease. Delayed patients had higher unadjusted rates of complete resection (93.7% vs. 91.9%, P = 0.032) and lower rates of emergency surgery (4.5% vs. 22.5%, P < 0.001). After adjustment, delay was not associated with a lower rate of complete resection (OR 1.18, 95% CI 0.90-1.55, P = 0.224), which was consistent in elective patients only (OR 0.94, 95% CI 0.69-1.27, P = 0.672). Longer delays were not associated with poorer outcomes. CONCLUSION: One in 15 colorectal cancer patients did not receive their planned operation during the first wave of COVID-19. Surgical delay did not appear to compromise resectability, raising the hypothesis that any reduction in long-term survival attributable to delays is likely to be due to micro-metastatic disease

Synthetic Studies Towards Prismanes - Rearrangements In Some Pentacyclic Precursors And X-Ray Crystal-Structure Of A Novel Heptacyclic Ether

In order to gain access to the heptacyclic tetraone 3, efforts were directed towards the utilisation of the major 'unwanted' [4 + 2]-adduct 11 of tetrachlorodimethoxycyclopentadiene and norbornenobenzoquinone. Epoxides derived from the diol and dimethoxy derivatives of the adduct 11 undergo facile Wagner-Meerwein rearrangement resulting in the required endo, syn, endo stereochemistry as well as methano-bridge functionalisation to deliver 18 and 24, respectively. However, intramolecular ether formation, occurring via the capture of carbocation intermediate with the transannularly poised oxygen functionality, is a more facile process. Attempts to cleave the ether linkage resulted in the formation of a novel transannularly cyclised twisted bowl shape heptacyclic compound 30 and its structure has been established through X-ray crystallography

Discriminatory protein binding by a library of 96 new affinity resins: A novel dye-affinity chromatography tool-kit

Initial acceptance of Cibacron Blue 3G-A (R) based matrices has made dye-ligand affinity chromatography an attractive proposition. This prompted the synthesis and search for new dye structures. A systematic library of 96 affinity resins was generated using novel analogs of Cibacron Blue 3G-A (R) and also by varying spacer lengths for immobilization. The library was tested in a batch binding and elution mode using seven different proteins - four Aspergillus enzymes namely, NADP-glutamate dehydrogenase, laccase, glutamine synthetase and arginase, bovine pancreatic trypsin and the two serum proteins human serum albumin and immunoglobulin G. Unique binding patterns were observed for each of them indicating that the library displayed discriminatory interactions. The significance of spacer length in the interaction with proteins was discernable. Trypsin interacted best with affinity resins that had no spacer. It was possible to resolve IgG and HSA from a mixture using a combination of resins. There was a good spread of HSA binding capacity in the 96 affinity resins. While some showed better HSA binding capacity than the commercial CB3GA-based matrix, a few with lower capacity were also observed. Subsequent to an initial screen, one affinity resin (CR-017) could be used to enrich Aspergillus terreus NADP-GDH from crude cell extracts. The efficacy of this dye-affinity resin was rationalized by characterizing NADP-GDH inhibition kinetics with the corresponding free dye ligand. In the sum, the library provides a set of dye-ligand affinity matrices with a potential for use in high throughput screening for protein purification. (C) 200