iBB

Gold(III) bisdithiolate complexes have been shown as potential antimicrobial and antitumoral agents. In particular, the complex [Au(cdc)2]− (cdc=cyanodithioimido carbonate) displayed antimicrobial and outstanding antitumor activity against the ovarian cancer cells A2780 and A2780cisR, which are sensitive and resistant to cisplatin, respectively. Yet, poor water solubility may hinder its clinical use. Block copolymer micelles (BCMs) have demonstrated great potential as nanocarriers capable of delivering hydrophobic drugs, increasing their solubility and stability, and improving their bioavailability and circulation time in blood, with reduced side effects. In this work, published in the journal Pharmaceutics, the potential of BCM-[Au(cdc)2] as a novel drug-delivery system was addressed. The BCM-[Au(cdc)2] micelles were prepared with a loading efficiency of 64.6% and a loading content of 35.3 mg [Au(cdc)2]−/gBCM, they were also homogenous and good candidates for drug delivery. Cytotoxic activity studies against A2780/A2780cisR cells showed that BCM-[Au(cdc)2] maintained relevant cytotoxic activity comparable to the cytotoxicity observed for the same concentration of gold complexes. The Au uptake in A2780 cells, determined by PIXE, was ca. 17% higher for BCMs-[Au(cdc)2] compared to [Au(cdc)2]−. The BCMs-[Au(cdc)2] presented also antimicrobial activity against S. aureus Newman and C. glabrata CBS138. In summary this paper shows the results of the potential of BCM-[Au(cdc)2] for drug delivery and its promising anticancer and antimicrobial activities.

This study was led by Dr. Célia Fernandes and Dr. Fernanda Marques, both from Centro de Ciências e Tecnologias Nucleares, Instituto Superior Técnico. The iBB members Teresa Pinheiro, Jorge Leitão and Sílvia Sousa collaborated in this work performing the cellular uptake analysis and the antimicrobial activities of BCMs.

RNA-binding proteins (RBPs) are important regulators of cellular functions, playing critical roles on the survival of bacteria and in the case of pathogens, on their interaction with the host. RBPs are involved in transcriptional, post-transcriptional, and translational processes. However, except for model organisms like Escherichia coli, there is little information about the identification or characterization of RBPs in other bacteria, namely in members of the Burkholderia cepacia complex (Bcc). Bcc is a group of bacterial species associated with a poor clinical prognosis in cystic fibrosis patients. These species have some of the largest bacterial genomes, and except for the presence of two-distinct Hfq-like proteins, their RBP repertoire has not been analyzed so far. Using in silico approaches, we identified 186 conventional putative RBPs in Burkholderia cenocepacia J2315, an epidemic and multidrug resistant pathogen of cystic fibrosis patients. In a work recently published in the journal GENES, Joana Feliciano, António Seixas, Tiago Pita and Jorge H. Leitão described the comparative genomics and phylogenetic analysis of RBPs present in multiple copies and predicted to play a role in transcription, protein synthesis, and RNA decay in Bcc bacteria. The analysis suggest the existence, in B. cenocepacia and in other Bcc bacteria, of some extra and unexplored functions for the mentioned RBPs, as well as of alternative mechanisms involved in RNA regulation and metabolism in these bacteria.

The synthesis of hydroxide [Ag(OH)L] (L =I VL, VL, VIL, VIIL), oxide [{AgL}2}(μ-O)] (L = IL, IIL, IIIL, VL, VIL) or chloride [AgIIL]Cl, [Ag(VIL)2]Cl complexes, obtained from reactions of mono- or bicamphorimine derivatives with Ag(OAc) or AgCl, appeared in a recent publication on the journal Antibiotics, resulting from a collaborative research involving the iBB members Jorge H. Leitão, Nuno Mira, Ana Rego and Sílvia Sousa; M. Fernanda Carvalho from CQE; and Fernanda Marques from C2TN. The new complexes were characterized by spectroscopic (NMR, FTIR) and elemental analysis. X-ray photoelectron spectroscopy (XPS), ESI mass spectra and conductivity measurements were undertaken to corroborate formulations. The antimicrobial activity of complexes and some ligands were evaluated towards Candida albicans and Candida glabrata, and strains of the bacterial species Escherichia coli, Burkholderia contaminans, Pseudomonas aeruginosa and Staphylococcus aureus based on the Minimum Inhibitory Concentrations (MIC). A significant feature of these redesigned complexes is their ability to sensitize C. albicans, a trait that was not found for the previously investigated [Ag(NO3)L] complexes. The MIC values of the complexes towards bacteria were in the range of those of [Ag(NO3)L] and well above those of the precursors Ag(OAc) or AgCl. The activity of the complexes towards normal fibroblasts V79 was evaluated by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay. Results showed that the complexes have a significant cytotoxicity.

The synthesis, characterization and antibacterial activity of eleven new complexes of general formula [Ag(NO3)(L-Y)2] corresponding to Ag(I) camphorimine complexes and a camphor sulfonylimine complex were reported in a recent publication in Journal of Inorganic Biochemistry, resulting from collaborative work of JH Leitão from BSRG and MF Carvalho from CQE. The study was performed to gain insights into structure/antimicrobial activity relationships. The lipophilicity and polarity which are important parameters concerning the biological activity of the complexes are also high dependent of the characteristics of the camphor ligands. The redox properties of the complexes studied by cyclic voltammetry showed that their reduction potentials are essentially independent of their electronic and steric properties. The antibacterial activity of all the complexes, against Gram-positive (S. aureus Newman) and Gram-negative (Escherichia coli ATCC25922, Pseudomonas aeruginosa 477, Burkholderia contaminans IST408) strains was evaluated through calculation of MIC values. Results show that complexes with camphor imine ligands that combine high lipophilicity with low dipolar moment exhibit enhanced antibacterial activity. The ability to establish hydrogen bonding emerged as an important contribution to the antibacterial activity of the camphor sulphonylimine complex.

Cystic fibrosis (CF) is the most life-limiting autosomal recessive disorder in Caucasians, with chronic bacterial airway infections representing the major cause of early decease. Pseudomonas aeruginosa and bacteria from the Burkholderia cepacia complex (Bcc) are the leading pathogens of CF patients’ airways. A wide array of virulence factors is responsible for infections caused by these bacteria, which have tightly regulated responses to the host environment. Small noncoding RNAs (sRNAs) are major regulatory molecules in these bacteria. Several approaches have been developed to study P. aeruginosa sRNAs, many of which are involved in the virulence. On the other hand, the knowledge on Bcc sRNAs remains far behind. In a review published on International Journal of Molecular Sciences, Tiago Pita, Joana Feliciano and Jorge H. Leitão from BSRG-iBB updated the knowledge on characterized sRNAs involved in P. aeruginosa virulence, compiled data so far achieved on sRNAs from the Bcc and discuss their possible roles on bacteria virulence.

 Bacteria of the Burkholderia cepacia complex (Bcc) remain an important cause of morbidity and mortality among patients suffering from cystic fibrosis. Eradication of these pathogens by antimicrobial therapy often fails, highlighting the need to develop novel strategies to eradicate infections. Vaccines are attractive since they can confer protection to particularly vulnerable patients, as is the case of cystic fibrosis patients. Several studies have identified specific virulence factors and proteins as potential subunit vaccine candidates. So far, no vaccine is available to protect from Bcc infections. In a recent publication in the journal Vaccines, iBB-BSRG researchers Sílvia Sousa, António M. Seixas and Jorge H. Leitão review the most promising postgenomic approaches and selected web tools available to speed up the identification of immunogenic proteins with the potential of conferring protection against Bcc infections.

Organized at Instituto Superior Técnico by Isabel Sá-Correia and Jorge Leitão, a joint workshop of two FCT-funded doctoral programs in which  DBE and iBB participate or direct, AEM-Applied and Environmental Microbiology and BIOTECnico- Biotechnology and Biosciences, took place at the Zoom platform on February 15 (dedicated to AEM) and 16 (dedicated to BIOTECnico). The external supervisory committee (ESC) of the two programs were present and participated very actively. They are highly renowned academics in the areas of the PhD programs: Tom Coyne from Gent University, William de Vos from Wageningen University, Jack Pronk from Delft University, Prof Steve Oliver, University of  Cambridge, Michael Hoare, University College London. 

PhD students and alumni of both programs were present and program that we set up via Zoom allowed us to demonstrate the work carried out in terms of advanced training in the framework of BIOTECnico and AEM programs. The ESCs were very impressed about the way how “the two fabulous programs were designed and managed and built a culture that encourages PhD candidates to take initiative, grow and prosper.” Congratulations to the directive boards of both programs, the students, the alumni, and their supervisors and collaborators.

In a paper recently published in the journal Antibiotics, a series of cyclam- and cyclen-derived salts were designed and studied as antibacterials by the research groups headed by JH Leitão (BSRG) and AM Martins and LG Alves (CQE). The compounds were designed specifically to gain insights into their structure and antibacterial activity towards Staphylococcus aureus and Escherichia coli, used respectively, as Gram-positive and Gram-negative model organisms. The newly synthesized compounds are monosubstituted and trans-disubstituted tetraazamacrocycles that display benzyl, methylbenzyl, trifluoromethylbenzyl, or trifluoroethylbenzyl substituents appended on the nitrogen atoms of the macrocyclic ring. The results obtained show that the chemical nature, polarity, and substitution patterns of the benzyl groups, as well as the number of pendant arms, are critical parameters for the antibacterial activity of the cyclam-based salts. The paper was published as part of the journal Special Issue New Insights into Antibacterial Compounds: From Synthesis and Discovery to Molecular Mechanisms of Action, guest-edited by Jorge H. Leitão.

Jorge H. Leitão from BSRG-iBB is guest-editing a special issue for the open-access journal "Antibiotics” entitled “New insights into antibacterial compounds: from synthesis and discovery to molecular mechanisms of action” . This issue aims to gather papers describing novel antibiotics, originating form chemical synthesis, repositioning of existent drugs, or from natural sources like plant extracts, herbs and spices. Papers on the description of novel structures, discovery of novel targets and mechanisms of action, as well as on the use of Omics approaches in the field of novel antibiotics discovery and characterization are also welcome.

Bacteria of the Burkholderia cepacia complex (Bcc) are ubiquitous multidrug resistant organisms and opportunistic pathogens capable of causing life threatening lung infections among cystic fibrosis (CF) patients. No effective therapies are available to eradicate Bcc bacteria from CF patients, as these organisms are inherently resistant to most clinically available antimicrobials. In a collaborative research work headed by BSRG members JH Leitão and SA Sousa, in collaboration with the CBMA (UMinho) team headed by Pedro Santos, and the Cystic Fibrosis center (HSM) headed by MDs C Barreto and L Pereira, an immunoproteomics approach was used to identify Bcc proteins that stimulate the humoral immune response of the CF host. The work was published in the journal New Biotechnology and reports for the first time the identification of 19 proteins as immunogenic. Ten proteins were predicted as extracytoplasmic, 9 of them being conserved in Bcc genomes. The immunogenic Bcc extracytoplasmic proteins are potential targets for development of novel therapeutic strategies and diagnostic tools to protect patients against the onset of chronic Bcc lung infections.

The emergence of bacterial resistance to available antimicrobials has prompted the search for novel antibacterial compounds to overcome this public health problem. Metal-based complexes have been much less explored than organic compounds as antimicrobials, leading to investigations of the antimicrobial properties of selected complexes in which silver may occupy the frontline due to its use as medicine since ancient times. Like silver, camphor has also long been used for medicinal purposes. However, in both cases, limited information exists concerning the mechanisms of their antimicrobial action. In a recent collaborative work by the research group headed by Jorge H. Leitão from BSRG/iBB and M. Fernanda Carvalho from CQE, the present knowledge of the antimicrobial properties of camphor-derived silver complexes is reviewed, focusing on recent research on the synthesis and antimicrobial properties of complexes based on silver and camphor imines. Selected examples of the structure and antimicrobial activity relationships of ligands studied so far are presented, showing the potential of silver camphorimine complexes as novel antimicrobials. The review was published in the journal Antibiotics.